The total synthesis of glucosepane and related chemical reactions, compounds and compositions and obtained therefrom and methods of treatment

ABSTRACT

Glucosepane is a structurally complex protein post-translational modification (PTM) believed to exist in all living organisms. Research in humans suggests that glucosepane plays a critical role in the pathophysiology of both diabetes and human aging; yet comprehensive biological investigations of this metabolite have been greatly hindered by a scarcity of chemically homogeneous material available for study. Glucosepane possesses a unique chemical structure that incorporates a surprising, never-before-prepared non-aromatic tautomer of imidazole (hereafter termed an “iso-imidazole”), rendering it a challenging target for chemical synthesis. In this application, the inventors report the first total synthesis of glucosepane, enabled by the development of a novel one-pot method for preparation of the iso-imidazole core. The synthesis of the present invention is concise (8-steps starting from commercial materials), convergent, high-yielding (12% overall), and enantioselective. These results should prove useful to the art and practice of heterocyclic chemistry, and critical for the study of glucosepane and its role in human health and disease, especially the treatment of diabetic disorders or its impact on aging processes. Methods of synthesis, compounds obtained therefrom, pharmaceutical compositions and methods of treatment provide embodiments of the present invention.

RELATED APPLICATIONS

This application claims the benefit of support of United Statesprovisional application no. U.S. 62/232,626, entitled “The TotalSynthesis of Glucosepane and Related Chemical Reactions”, filed Sep. 25,2015, the entire contents of which application is incorporated byreference herein.

FIELD OF THE INVENTION

The present invention is directed to a novel, highly efficient totalsynthesis of glucosepane and related derivatives and chemical reactionswhich make this synthesis possible. In particular, methods ofintroducing an iso-imidazole moiety onto a substituted oxo-azepineprecursor to provide glucosepane and derivatives thereof are disclosed.Additional embodiments of the present invention include compounds (fortheir biological activity and/or their use as synthetic intermediates,pharmaceutical compositions and methods of treatment as otherwisedescribed herein.

BACKGROUND OF THE INVENTION

Post-translation modifications (PTMs) of proteins are responsible for ahost of critical functions, ranging from accelerating protein folding tomediating protein-protein interactions.⁽¹⁾ Protein “glycation” is anon-enzymatic process for PTM formation wherein protein side-chainsreact spontaneously with open chain tautomers of carbohydrates. Mountingevidence suggests that protein glycation adducts, also called “advancedglycation end-products” or “AGEs”, are critically involved in bothhealthy and disease processes, including inflammation, diabetes, cancer,and normal human aging.^((2, 3)) Notably, AGEs often possess highlycomplex chemical structures, impeding their detailed chemical andbiological characterization.⁽⁴⁾

Glucosepane (1) is an important member of the AGE family that is bothbiologically and chemically significant (See FIG. 1). The molecule isformed as a “crosslink” from reaction sequences between arginine andlysine side-chains and one equivalent of hexose carbohydrate (mostcommonly glucose). Glucosepane is present on long-lived plasma proteinsin the human body, such as collagen and lens crystallin,^((3, 5)) and isalso found in high levels in various dietary sources, especiallyalkali-treated baked goods.⁽⁶⁾ Researchers have speculated thatglucosepane is directly involved in the pathophysiology of variousconditions (e.g., diabetes, diabetes-related complications, and aging)due to patterns of glucosepane formation on disease-associated proteins.For example, analysis of skin biopsies obtained through the DiabetesControl and Complications Trial (DCCT) has determined that increases inskin glucosepane levels represent a significant, independent risk factorfor the onset of diabetic nephropathy, retinopathy, andneuropathy.^((3, 7)) Additional studies have demonstrated thatnon-enzymatic glucosepane crosslinks outnumber enzyme-catalyzedcrosslinks in human collagen in people over 65 years of age. By age 100,glucosepane levels reach 2 nmol/mg collagen, which is almost ten timesnormal levels, whereas levels in diabetic patients can achieve up totwenty times those in healthy controls.^((9, 10))

Several mechanisms have been proposed for glucosepane's involvement indisease complications. For example, researchers have hypothesized thatglucosepane modification can decrease protein turnover rate and impairthe renewal of damaged proteins. Glucosepane crosslinks may also beresponsible for reported age- and diabetes-related decreases in collagendigestibility.^((3),(11, 12)(13)) Others have speculated thatglucosepane-induced Arg modification can decrease the number of integrinbinding sites in collagen, causing endothelial cell apoptosis,extracellular matrix deposition, and basement membrane thickening.(14)Glucosepane may also serve as ligand for pattern recognition receptorssuch as RAGE,⁽¹⁵⁾ leading to chronic inflammation, or as a neoepitopethat drives the breaking of self-tolerance against modifiedextracellular matrix proteins, serving as a trigger for the induction ofautoimmune processes. Finally, due to high levels of glucosepane andother AGEs in the human diet, it has been suggested that these materialsmay function as uremic toxins, leading to complications in the settingof renal failure.(16)

Despite glucosepane's health implications, biological investigationshave been hampered by a scarcity of chemically homogeneous materialavailable for study. Its complex non-enzymatic biosynthesis involvesserial tautomerizations of Amadori adduct 4 to provide glucosone 3 (aprocess termed “carbonyl mobility”, FIG. 1B). During this process, eachstereocenter undergoes epimerization, and therefore the glucosepane coreexists in nature as a mixture of all eight possiblediastereomers.^((3),(17)) These stereoisomers can only bechromatographically resolved into four binary mixtures, each containingtwo spectroscopically indistinguishable diastereomers with the samerelative configuration at the 6, 7, and 8a positions, but oppositeabsolute configurations with respect to the enantiomerically purebackbone amino acids.⁽¹⁷⁾ Despite significant effort, purification ofstereochemically homogeneous glucosepane from biological samples hasproven impossible. It is therefore unknown which of the eightstereoisomers is the most prevalent in vivo. Furthermore, these binarydiastereomeric mixtures can only be isolated in low yields (0.2-1.4%)following model reactions between lysine, arginine and glucose, andextensive chromatographic purification.^((17, 18)) Importantly, becauseof these difficulties in purification, antibody reagents to enablebiological detection of glucosepane in unprocessed tissue preparationsare unavailable. To our knowledge, therefore, all publishedinvestigations into glucosepane biology have relied upon time-consumingextraction protocols, involving exhaustive enzymatic hydrolysis followedby HPLC purification. The development of synthetic routes towardchemically-defined glucosepane constructs represents an essential nextstep toward understanding the roles that this compound plays in humanhealth and disease, and also toward the identification of noveltherapeutic and/or diagnostic agents.

Glucosepane presents a deceptively challenging synthetic target due toits high density of heteroatoms, the presence of a stereogenic polyolmotif incorporated within a fused hetero-bicyclic topology, anepimerizable stereocenter at C-8a, and perhaps most notably, thepresence of an arginine-derived iso-imidazole at its core. Indeed, atfirst glance, one would expect glucosepane to tautomerize spontaneouslyto the corresponding aromatic imidazole (FIG. 1A); however, reportedstructural assignments of the iso-imidazole in glucosepane areconsistent with one- and two-dimensional NMR data reported by Ledererand colleagues.⁽¹⁷⁾ Furthermore, because glucosepane forms naturally asa protein adduct (not as the free bis-amino acid crosslink), any usefulsynthesis needs to be compatible with glucosepane incorporation intopeptides. Also, because glucosepane is formed naturally as a mixture ofall eight possible diastereomers, synthetic efforts targeting bothenantio- and diastereomerically pure material are essential for detailedbiochemical study.

BRIEF DESCRIPTION OF THE INVENTION

In one embodiment, the invention provides novel processes for makingsubstantially pure glucosepane and substantially pure glucosepanederivatives in relatively large yields through syntheses that employsignificantly fewer steps than known techniques.

In another embodiment, the invention provides novel processes for makingvarious intermediates useful in the manufacture ofpharmaceutically-active ingredients, including substantially pureglucosepane and substantially pure glucosepane derivatives.

In still another embodiment, the invention provides various novelcompounds and intermediates useful in the manufacture ofpharmaceutically-active ingredients, including substantially pureglucosepane and substantially pure glucosepane derivatives.

In still another embodiment, the invention provides methods of treating,inhibiting or reducing the likelihood of diabetes or a diabetes relateddisorder/secondary condition or inhibiting aging processes of the bodyadministering either substantially pure glucosepane or a glucosepanederivative to a subject who suffers from, or who is at risk ofdeveloping, a disease state, disorder or aging processes as set forthabove.

The present invention is directed to a method of synthesizingisoimidazoles, especially glucosepane according to the method which isset forth in scheme 3 herein (FIG. 5). The method comprises reacting anazepin-one compound with a semicarbazone to provide an intermediatecompound (where R and X are as described generically below):

which can be reacted with a cyclizing agent (preferably trimethylsilylchloride in solvent) to provide the compound:

Where Y′ is H,

or a non-salt (free form), an alternative salt form or stereoisomerthereof.

Compound 28A can be further manipulated to provide compounds accordingto the present invention including the preferred glucosepane or ananalog (in preferred embodiments, R is obtained from a lysine oromithine amine acid, X is obtained from an ornithine or lysine aminoacid and Y′ is H.

Thus, in one embodiment, in the present method an azepin-one compoundaccording the chemical structure (14)

where R is a C₁-C₁₂ optionally substituted hydrocarbon group (preferablyan optionally substituted alkyl or aryl group) or a heterocyclic group,preferably a heteroaryl group (in preferred aspects N—CH₂—R of theazepine ring is a lysine or ornithine moiety with the amine group formedfrom the distal amino groups of the side chain of the amino acid) isreacted according to scheme 3, FIG. 5 with a semicarbazone compound(which may be a salt form, depending on the conditions used) accordingto the chemical structure:

Where X is an optionally substituted S-alkyl (preferably, C₁-C₇ alkyl,preferably, S-Me or an amino acid group obtained from cysteine), anoptionally substituted S-aryl or an optionally substitutedS-heterocyclyl (preferably, S-heteroaryl), an optionally substitutedO-alkyl (preferably, C₁-C₇ alkyl, preferably. O-Me or an amino acidgroup obtained from serine), an optionally substituted O-aryl (or anoptionally substituted O-heterocyclyl (preferably, O-heteroaryl), a NRR² group where R¹ and R² are each independently H (often. R² is H), anoptionally substituted alkyl group (preferably, an optionallysubstituted C₁-C₁₂, preferably C₁-C₇ alkyl group including a C₃ or C₄alkylene amino acid group obtained from lysine or ornithine, wherein theamino group and/or the carboxylic acid group is preferably protected),an optionally substituted aryl group or an optionally substitutedheterocyclyl (preferably an optionally substituted heteroaryl), or (Xis) an amino acid group preferably obtained from a D- or L-amino acidaccording to the chemical structure:

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline) and R³ is a side chain derived from an amino acidpreferably selected from the group consisting of alanine (methyl),arginine (propyleneguanidine), asparagine (methylenecarboxyamide),aspartic acid (ethanoic acid), cysteine (thiol, reduced or oxidizeddi-thiol), glutamine (ethylcarboxyamide), glutamic acid (propanoicacid), glycine (H), histidine (methyleneimidazole), isoleucine(1-methylpropane), leucine (2-methylpropane), lysine (butyleneamine),ornithine (propyleneamine), methionine (ethylmethylthioether),phenylalanine (benzyl), proline (R³ forms a cyclic ring with R_(a) andthe adjacent nitrogen group to form a pyrrolidine group),hydroxyproline, serine (methanol), threonine (ethanol, 1-hydroxyethane),tryptophan (methyleneindole), tyrosine (methylene phenol) or valine(isopropyl), where the R³ side chain is optionally protected. PreferablyX is an optionally substituted S—(C₁-C₁₂) alkyl (more preferably, S-Me),O—(C₁-C₁₂) alkyl or a NH—R¹ group where R¹ is an optionally substitutedC₁-C₁₂, preferably a C₁-C₁₀ alkyl group (preferably, NHR¹ is anornithine or lysine moiety, which contains protecting groups) to providecompound 24A:

where R and X are the same as above;Compound 24A is further reacted with trimethylsilyl chloride (TMSCl) inthe presence of solvent (preferably chloroform, methylene chloride) atelevated temperature (generally, above room temperature and often at thereflux temperature of the solvent used) to provide compound 28A

or an alternative pharmaceutical salt or neutral i.e. non-salt compoundor stereoisomer thereof;Where Y′ is H and R and X are the same as above, preferably in one pot(compound 14 can be converted to compound 28A in a single pot andcompound 24A can be converted to compound 28A in a single pot); andoptionally, deprotecting the protected compound, which can be performedin the same pot or separated prior to deprotection. In this method,preferably N—CH₂—R of the azepine ring is a lysine or ornithine moietywith protecting groups on the amine and optionally the carboxylic acidmoieties of R and X is NHR¹ as an ornithine or lysine moiety, also withoptional protecting groups on the amine and optionally the carboxylicacid moieties on the ornithine and lysine moieties.

In certain embodiments of the present invention, in compound 28A,including where X is S-alkyl, even more preferably S-Me, Y′ can beconverted to a hydroxyl group using SiO₂ (FIG. 5, Scheme 3, compound 26is converted to compound 27 using SiO₂) or triethylamine inacetonitrile/water (FIG. 6 Scheme 3A) and the intermediate compound (Y′is OH, X is as described above, preferably S-Me of FIG. 5, Scheme 3) canbe deprotected or further reacted with an amine (including an amino acidderivative), preferably a protected ornithine or lysine amino acid (theamine on the side chain is unprotected, with the amine group andoptionally the carboxylic acid of the amino acid group being protected,preferably with a Cbz or other protecting group) to introduce the amineinto the compound as a substituent on the isoimidazole moiety (seecompound 28A above or compound 28 of FIG. 5, Scheme 3), followed byreduction, preferably with a borohydride (e.g. Na(OAc)₃BH) or otherreducing agent (to convert Y′ as OH to H) to produce a compound where Xis as described above, preferably an amine, more preferably a protectedorninthine or lysine group (protecting group at least on the amine)which compound may be subsequently deprotected to provide compound 28A,

Y′ is H, NCH₂R is a lysine or ornithine (preferably lysine) moiety and Xis a lysine or ornithine (preferably ornithine) moiety. The protectinggroups may be removed pursuant to the selectivity of the protectinggroup to removal as described herein, consistent with otherwisemaintaining the integrity of the chemistry of the compound.

In preferred aspects, the formation of compound 28A from compound 14occurs in a single pot reaction in high yield (often greater than 50%,55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% and higher).

In certain preferred aspects of the present invention, compound 14(where N—CH₂—R of the azepine ring is a lysine or ornithine moiety,preferably a lysine moiety, with the amine group formed from the distalamino groups of the side chain of the amino acid) is reacted with asemicarbazone of the chemical structure:

Where X is an optionally substituted S-alkyl (preferably, C₁-C₇ alkyl,preferably, S-Me or an amino acid group obtained from cysteine), anoptionally substituted S-aryl or an optionally substitutedS-heterocyclyl (preferably, S-heteroaryl), an optionally substitutedO-alkyl (preferably, C₁-C₇ alkyl, preferably, O-Me or an amino acidgroup obtained from serine), an optionally substituted O-aryl (or anoptionally substituted O-heterocyclyl (preferably, S-heteroaryl), aNR¹R² group where R¹ and R² are each independently H (often, R² is H),an optionally substituted alkyl group (preferably, a C₁-C₇ alkyl groupor an amino acid group obtained from lysine or ornithine), an optionallysubstituted aryl group or an optionally substituted heterocyclyl(preferably an optionally substituted heteroaryl), or (X is) an aminoacid group preferably obtained from a D- or L-amino acid according tothe chemical structure:

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline or hydroxyl proline) and R³ is a side chain derived from anamino acid preferably selected from the group consisting of alanine(methyl), arginine (propyleneguanidine), asparagine(methylenecarboxyamide), aspartic acid (ethanoic acid), cysteine (thiol,reduced or oxidized di-thiol), glutamine (ethylcarboxyamide), glutamicacid (propanoic acid), glycine (H), histidine (methyleneimidazole),isoleucine (1-methylpropane), leucine (2-methylpropane), lysine(butyleneamine), methionine (ethylmethylthioether), phenylalanine(benzyl), proline (R³ forms a cyclic ring with R_(a) and the adjacentnitrogen group to form a pyrrolidine group), hydroxyproline, serine(methanol), threonine (ethanol, 1-hydroxyethane), tryptophan(methyleneindole), tyrosine (methylene phenol) or valine (isopropyl)(where the R³ side chain is optionally protected). Preferably X is anoptionally substituted S-alkyl (preferably. S-Me) or a NH—R¹ group whereR¹ is an ornithine or lysine moiety, which contains protecting groups onthe amine and carboxylic acid groups) react to form the compound 24A

where R and X are the same as above;Compound 24A is further reacted with trimethylsilyl chloride (TMSCl) inthe presence of solvent (preferably chloroform) at elevated temperature(generally, above room temperature and often at the reflux temperatureof the solvent used) to provide compound 28A:

Or an non-salt form, alternative salt form or stereoisomer thereof,Where Y′ is H and R and X are the same as above (preferably N—CH₂—R ofthe azepine ring is a lysine or ornithine moiety and X is NHR¹ as anornithine or lysine moiety as described herein); and optionally,deprotecting the protected compound. In preferred aspects, the formationof compound 28A from compound 14 occurs in a single pot reaction in highyield (often greater than 50%, 55%, 60%, 70%, 80% and higher). Note thatcompound 28A may also be in a non-salt or alternative salt form or asone of a number of stereoisomers, including a diastereomer orenantiomer). Compound 28A may be converted such that Y′ is OH byreacting compound 28A with SiO2 or triethylamine in aqueous solvent(aqueous acetonitrile) to convert Y′ as H to OH. When Y′ is OH, thehydroxyl group may be converted back to a hydrogen group by reducing thehydroxyl to hydrogen using a reducing agent, preferably a borohydridereducing agent (e.g. Na(OAc)₃BH). In certain embodiments (preferablywhere X is S-alkyl, especially SMe), X may also be substituted by anamine, preferably an amino acid, including an ornithing or lysine group(attachment is on the distal amine of the side chain) as otherwisedescribed herein.

In another embodiment (see FIG. 3, Scheme 1 and FIG. 5, Scheme 3), thepresent invention relates to a total synthesis of glucosepane from thereadily available epoxide starting material 8

Comprising exposing compound 8 to a protected amine, preferably aprotected amino acid (preferably a lysine or ornithine derivative)according to the chemical structure:

BL-NH—CH₂R

where R is an optionally substituted C₁-C₁₂ hydrocarbon group(preferably an optionally substituted alkyl or aryl group, preferablyphenyl) or an optionally substituted heterocycle group (preferably, aheteroaryl group), preferably R is an alkylene amino acid group(preferably from lysine or ornithine which is protected on the amine andcarboxylic acid groups of the amino acid)and BL is a protecting group (preferably, a Dod blocking/protectinggroup or other protecting group group which does not impair or preventthe amine group reacting with the epoxide of compound 8) in the presenceof solvent (preferably ethanol, isopropanol) at elevated temperature ina first step followed by strong acid, preferably a trifluoroacetic acidsolution (e.g. 5%) in solvent (preferably, methylene chloride,chloroform) in the presence of anisole and a hydrosilane (preferablyiPr₃SiH or other trialkylhydrosilane) to provide compound 10:

Where R is the same as described above:Compound 10 is thereafter exposed to aqueous acid (preferably 70%aqueous acetic acid or other organic acid at elevated temperature) toprovide compound 11 (FIG. 3, scheme 1), which spontaneously undergoesamadori rearrangement and intramolecular trapping to provide compound13:

which is reacted with 2,2-dimethoxypropane (DMP) in the presence ofpyridinium p-toluenesulfonate (PPTS) in solvent (toluene) to providedihydroxy protected compound 14, which can then be reacted with thesemicarbazone as described above (especially compound 22 or 23 of FIG.5, Scheme 3 A., followed by reaction with trimethylsilylchloride (TMSCl)at elevated temperature and deprotection of any protecting groups toprovide compound 28A

Where X and R are the same as above and Y′ is H (or the non-salt oralternative salt thereof or a stereoisomer thereof).

In preferred aspects, where the semicarbazone is compound 22 or 23 (FIG.5, Scheme 3A.), X is SMe, HN-Orn or HN-Lys (the lysine is protected inan analogous manner to HN-Orn, depicted) and Y′ is H (or the free amineor alternative salt thereof) (see compounds 26 or 28 of FIG. 5, scheme3), which compounds can then be deprotected to provide the finaldeprotected compounds.

In certain preferred aspects, compound 26

is reacted with SiO2 or triethylamine in aqueous solvent (e.g.acetonitrile/water) to provide a hydroxyl substituted compound 27 (FIG.5, Scheme 3, and FIG. 6, scheme 3A, the bridge hydrogen is replaced witha hydroxyl group) which may be further reacted with a protectedornithine (or analogous lysine derivative) or other amine (which isoptionally protected) optionally in a weak base (e.g. triethylamine)followed by borohydride reduction (preferably Na(OAc₃)BH) to produce anornithine (or lysine/amine) compound 28 which is subsequently optionallydeprotected, for example using reduction or other conditions to removeany protecting groups to provide a final deprotected compound.

As discussed above, compound 26

is alternatively reacted with SiO₂ or in weak base (e.g. triethyl amine)in aqueous solvent (e.g. acetonile or other polar solvent) to providecompound 27 (the bridge hydrogen is converted to hydroxyl). Compound 27can be converted to compound 28 by reacting compound 27 with a protectedornithine, lysine or other amine followed by borohydride reduction ofthe bridge hydroxyl to hydrogen and subsequently deprotecting anyprotecting group(s) to provide the final deprotected compound (or itsnon-salt, alternative salt or a stereoisomer thereof).

A large number of glucosepane derivatives may be made by the samemethod. Note that compound 28A is in equilibrium with its free amine(i.e. non-salt form) and can be found in its free amine or as analternative salt form, as well as in a number of stereoisomeric forms.

In still another embodiment, the present invention is directed to amethod for synthesizing an optionally substituted imidazole from analdehyde or ketone of the general formula 1k:

Where Z¹ is H, an optionally substituted C₁-C₁₂ hydrocarbon group(preferably an alkyl, alkenyl or aryl group), a 3-20 membered(preferably, a 5- to 20-membered) heterocyclic group (preferably, aheteroaryl group), a NR¹R² group, a SR¹ or OR¹ group or together Z¹ andZ² link to form an optionally substituted 5- to 8-membered ring (withthe carbonyl and methylene group) which ring is carbocyclic orheterocyclic, including one or more unsaturated bonds, aryl orheteroaryl;Z² is H, an optionally substituted C₁-C₁₇ hydrocarbon group (preferablyan alkyl, alkenyl or aryl group), a 3-20 membered (preferably, a 5- to20-membered) heterocyclic group (preferably, a heteroaryl group) ortogether Z¹ and Z² are linked to form an optionally substituted 5- to8-membered ring (preferably, a 6- or 7-membered ring) which iscarbocyclic or heterocyclic, including aryl or heteroaryl;R¹ and R² are each independently absent (only one of R¹ and R² may beabsent), H, an optionally substituted C₁-C₆ alkyl, alkene or alkynegroup, an optionally substituted aryl or heterocyclic group (preferably,including a heteroaryl group) or NR¹R² is an optionally protected aminoacid group where R¹ is H or a C₁-C₃ alkyl group and R² is a grouppreferably obtained from a D- or L-amino acid according to the chemicalstructure:

where R³ is a side chain derived from an amino acid preferably selectedfrom the group consisting of alanine (methyl), arginine(propyleneguanidine), asparagine (methylenecarboxyamide), aspartic acid(ethanoic acid), cysteine (thiol, reduced or oxidized di-thiol),glutamine (ethylcarboxyamide), glutamic acid (propanoic acid), glycine(H), histidine (methyleneimidazole), isoleucine (1-methylpropane),leucine (2-methylpropane), lysine (butyleneamine), ornithine(propyleneamine), methionine (ethylmethylthioether), phenylalanine(benzyl), proline (R³ forms a cyclic ring with the adjacent nitrogengroup when R¹ is H to form a pyrrolidine group), hydroxyproline, serine(methanol), threonine (ethanol, 1-hydroxyethane), tryptophan(methyleneindole), tyrosine (methylene phenol) or valine (isopropyl),The method comprising reacting a compound of formula 1i with asemicarbazone compound of formula S1:

Where X¹ is an optionally substituted S-alkyl (preferably, C₁-C₇ alkyl,preferably, S-Me or an amino acid group obtained from cysteine), anoptionally substituted S-aryl or an optionally substitutedS-heterocyclyl (preferably, S-heteroaryl), an optionally substitutedO-alkyl (preferably, C₁-C₇ alkyl, preferably, O-Me or an amino acidgroup obtained from serine), an optionally substituted O-aryl (or anoptionally substituted O-heterocyclyl (preferably, S-heteroaryl), aNR¹R² group where R¹ and R² are each independently H (often, R² is H),an optionally substituted C₁-C₁₂ alkyl group (preferably, a C₁-C₇ alkylgroup or an amino acid group obtained from lysine or ornithine), anoptionally substituted aryl group or an optionally substitutedheterocyclyl (preferably an optionally substituted heteroaryl), or (Xis) an amino acid group preferably obtained from a D- or L-amino acid

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline) and R³ is a side chain derived from an amino acidpreferably selected from the group consisting of alanine (methyl),arginine (propyleneguanidine), asparagine (methylenecarboxyamide),aspartic acid (ethanoic acid), cysteine (thiol, reduced or oxidizeddi-thiol), glutamine (ethylcarboxyamide), glutamic acid (propanoicacid), glycine (H), histidine (methyleneimidazole), isoleucine(1-methylpropane), leucine (2-methylpropane), lysine (butyleneamine),methionine (ethylmethylthioether), phenylalanine (benzyl), proline (R³forms a cyclic ring with R_(a) and the adjacent nitrogen group to form apyrrolidine group), hydroxyproline, serine (methanol), threonine(ethanol, 1-hydroxyethane), tryptophan (methyleneindole), tyrosine(methylene phenol) or valine (isopropyl) (where the R³ side chain and/orthe carboxylic acid group is optionally protected) to obtain a compoundof the formula 1ia:

or its salt (preferably a pharmaceutically acceptable salt)where Z¹, Z² and X¹ are the same as above,which is thereafter reacted with trimethylsilyl chloride in solvent(e.g. chloroform) at elevated temperature to obtain the compound 1i

(as a hydrochloride salt, non-salt or alternative salt form),

Where Y′ is H, and

Wherein said compound is optionally and preferably deprotected.

In further embodiments, Y′ in the above imidazole may be converted to ahydroxyl group in SiO₂ or weak base (triethylene amine) in aqueoussolvent (preferably acetonitrile). In addition, the X¹ substituent (e.g.S-alkyl) may be converted to another substituent (for example,especially with an amine).

The present invention is also directed to compounds according to thechemical structure (or a salt form):

where R is a C₁-C₁₂ optionally substituted hydrocarbon group (preferablyan optionally substituted alkyl or aryl group) or a heterocyclic group,preferably a heteroaryl group (in preferred aspects N—CH₂—R of theazepine ring is a lysine or ornithine moiety with the amine group formedfrom the distal amino groups of the side chain of the amino acid); andX is optionally substituted S-alkyl (preferably, C₁-C₇ alkyl,preferably, S-Me or an amino acid group obtained from cysteine), anoptionally substituted S-aryl or an optionally substitutedS-heterocyclyl (preferably, S-heteroaryl), an optionally substitutedO-alkyl (preferably, C₁-C₇ alkyl, preferably, O-Me or an amino acidgroup obtained from serine), an optionally substituted O-aryl (or anoptionally substituted O-heterocyclyl (preferably, S-heteroaryl), aNR¹R² group where R¹ and R² are each independently H (often, R² is H),an optionally substituted alkyl group (preferably, a C₁-C₇ alkyl groupor an amino acid group obtained from lysine or ornithine), an optionallysubstituted aryl group or an optionally substituted heterocyclyl(preferably an optionally substituted heteroaryl), or (X is) an aminoacid group preferably obtained from a D- or L-amino acid according tothe chemical structure:

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline) and R³ is a side chain derived from an amino acidpreferably selected from the group consisting of alanine (methyl),arginine (propyleneguanidine), asparagine (methylenecarboxyamide),aspartic acid (ethanoic acid), cysteine (thiol, reduced or oxidizeddi-thiol), glutamine (ethylcarboxyamide), glutamic acid (propanoicacid), glycine (H), histidine (methyleneimidazole), isoleucine(1-methylpropane), leucine (2-methylpropane), lysine (butyleneamine),methionine (ethylmethylthioether), phenylalanine (benzyl), proline (R³forms a cyclic ring with R_(a) and the adjacent nitrogen group to form apyrrolidine group), hydroxyproline, serine (methanol), threonine(ethanol, 1-hydroxyethane), tryptophan (methyleneindole), tyrosine(methylene phenol) or valine (isopropyl) (where the R³ side chain isoptionally protected). Preferably X is an optionally substitutedS—(C₁-C₁₂) alkyl (preferably, S-Me), O—(C₁-C₁₂) alkyl or a NH—R¹ groupwhere R¹ is an optionally substituted C₁-C₁₂, preferably a C₁-C₀₁ alkylgroup (preferably, NHR¹ is an ornithine or lysine moiety,or a salt form) especially including a pharmaceutically acceptable saltform), stereoisomer (including a diastereomer or enantiomer), solvate orpolymorph thereof.

In another embodiment, the present invention is directed to compoundsaccording to the chemical structure:

Or a non-salt, alternative salt form or stereoisomer thereof,Where R is a C₁-C₁₂ optionally substituted hydrocarbon group (preferablyan optionally substituted alkyl or aryl group) or a heterocyclic group,preferably a heteroaryl group (in preferred aspects N—CH₂—R of theazepine ring is a lysine or ornithine moiety with the amine group formedfrom the distal amino groups of the side chain of the amino acid);X is optionally substituted S-alkyl (preferably, C₁-C₇ alkyl,preferably, S-Me or an amino acid group obtained from cysteine), anoptionally substituted S-aryl or an optionally substitutedS-heterocyclyl (preferably, S-heteroaryl), an optionally substitutedO-alkyl (preferably, C₁-C₇ alkyl, preferably, O-Me or an amino acidgroup obtained from serine), an optionally substituted O-aryl (or anoptionally substituted O-heterocyclyl (preferably, S-heteroaryl), aNR¹R² group where R¹ and R are each independently H (often, R² is H), anoptionally substituted alkyl group (preferably, a C₁-C₇ alkyl group oran amino acid group obtained from lysine or ornithine), an optionallysubstituted aryl group or an optionally substituted heterocyclyl(preferably an optionally substituted heteroaryl), or (X is) an aminoacid group preferably obtained from a D- or L-amino acid according tothe chemical structure:

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline) and R³ is a side chain derived from an amino acidpreferably selected from the group consisting of alanine (methyl),arginine (propyleneguanidine), asparagine (methylenecarboxyamide),aspartic acid (ethanoic acid), cysteine (thiol, reduced or oxidizeddi-thiol), glutamine (ethylcarboxyamide), glutamic acid (propanoicacid), glycine (H), histidine (methyleneimidazole), isoleucine(1-methylpropane), leucine (2-methylpropane), lysine (butyleneamine),methionine (ethylmethylthioether), phenylalanine (benzyl), proline (R³forms a cyclic ring with R_(a) and the adjacent nitrogen group to form apyrrolidine group), hydroxyproline, serine (methanol), threonine(ethanol, 1-hydroxyethane), tryptophan (methyleneindole), tyrosine(methylene phenol) or valine (isopropyl) (where the R³ side chain isoptionally protected). Preferably X is an optionally substitutedS—(C₁-C₁₂) alkyl (preferably, S-Me), O—(C₁-C₁₂) alkyl or a NH—R¹ groupwhere R¹ is an optionally substituted C₁-C₁₂, preferably a C₁-C₁₀ alkylgroup; and

Y is H or OH;

with the proviso that X, R and Y′ do not form glucosepane ora non-salt or alternative salt form, stereoisomer, solvate or polymorphthereof.

In yet another embodiment, the present invention is directed tocompounds according to the chemical structure:

Where Z′ is H, an optionally substituted C₁-C₁₂ hydrocarbon group(preferably an alkyl, alkenyl or aryl group), a 3-20 membered(preferably, a 5- to 20-membered) heterocyclic group (preferably, aheteroaryl group), a NR¹R² group, a SR¹ or OR¹ group or together Z¹ andZ² link to form an optionally substituted 5- to 8-membered ring (withthe carbonyl and methylene group) which ring is carbocyclic orheterocyclic, including one or more unsaturated bonds, aryl orheteroaryl;Z² is H, an optionally substituted C₁-C₁₂ hydrocarbon group (preferablyan alkyl, alkenyl or aryl group), a 3-20 membered (preferably, a 5- to20-membered) heterocyclic group (preferably, a heteroaryl group) ortogether Z¹ and Z² are linked to form an optionally substituted 5- to8-membered ring (preferably, a 6- or 7-membered ring) which iscarbocyclic or heterocyclic, including aryl or heteroaryl;R¹ and R² are each independently absent, H, an optionally substitutedC₁-C₆ alkyl, alkene or alkyne group, an optionally substituted aryl orheterocyclic group (preferably, including a heteroaryl group) or NR¹R²is an optionally protected amino acid group where R¹ is H or a C₁-C₃alkyl group and R² is a group preferably obtained from a D- or L-aminoacid according to the chemical structure:

where R³ is a side chain derived from an amino acid preferably selectedfrom the group consisting of alanine (methyl), arginine(propyleneguanidine), asparagine (methylenecarboxyamide), aspartic acid(ethanoic acid), cysteine (thiol, reduced or oxidized di-thiol),glutamine (ethylcarboxyamide), glutamic acid (propanoic acid), glycine(H), histidine (methyleneimidazole), isoleucine (1-methylpropane),leucine (2-methylpropane), lysine (butyleneamine), ornithine(propyleneamine), methionine (ethylmethylthioether), phenylalanine(benzyl), proline (R³ forms a cyclic ring with R_(a) and the adjacentnitrogen group to form a pyrrolidine group), hydroxyproline, serine(methanol), threonine (ethanol, 1-hydroxyethane), tryptophan(methyleneindole), tyrosine (methylene phenol) or valine (isopropyl);andX¹ is an optionally substituted S-alkyl (preferably, C₁-C₇ alkyl,preferably, S-Me or an amino acid group obtained from cysteine), anoptionally substituted S-aryl or an optionally substitutedS-heterocyclyl (preferably, S-heteroaryl), an optionally substitutedO-alkyl (preferably, C₁-C₇ alkyl, preferably, O-Me or an amino acidgroup obtained from serine), an optionally substituted O-aryl (or anoptionally substituted O-heterocyclyl (preferably, S-heteroaryl) or aNR¹R² group where R¹ and R² are each independently H (often, R² is H),an optionally substituted alkyl group (preferably, a C₁-C₇ alkyl groupor an amino acid group obtained from lysine or ornithine), an optionallysubstituted aryl group or an optionally substituted heterocyclyl(preferably an optionally substituted heteroaryl), or (X is) an aminoacid group preferably obtained from a D- or L-amino acid according tothe chemical structure:

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline) and R³ is a side chain derived from an amino acidpreferably selected from the group consisting of alanine (methyl),arginine (propyleneguanidine), asparagine (methylenecarboxyamide),aspartic acid (ethanoic acid), cysteine (thiol, reduced or oxidizeddi-thiol), glutamine (ethylcarboxyamide), glutamic acid (propanoicacid), glycine (H), histidine (methyleneimidazole), isoleucine(1-methylpropane), leucine (2-methylpropane), lysine (butyleneamine),methionine (ethylmethylthioether), phenylalanine (benzyl), proline (R³forms a cyclic ring with R_(a) and the adjacent nitrogen group to form apyrrolidine group), hydroxyproline, serine (methanol), threonine(ethanol, 1-hydroxyethane), tryptophan (methyleneindole), tyrosine(methylene phenol) or valine (isopropyl) (where the R³ side chain isoptionally protected). Preferably X is an optionally substitutedS—(C₁-C₁₂) alkyl (preferably, S-Me), O—(C₁-C₁₂) alkyl or a NH—R¹ groupwhere R¹ is an optionally substituted C₁-C₁₂, preferably a C₁-C₁₀ alkylgroup (preferably, NHR¹ is an ornithine or lysine moiety,or a salt form) especially including a pharmaceutically acceptable saltform), stereoisomer (including a diastereomer or enantiomer), solvate orpolymorph thereof.

In still further embodiments, the present invention is directed tocompounds according to the chemical structure:

Where Z¹ is H, an optionally substituted C₁-C₁₂ hydrocarbon group(preferably an alkyl, alkenyl or aryl group), a 3-20 membered(preferably, a 5- to 20-membered) heterocyclic group (preferably, aheteroaryl group), a NR¹R² group, a SR¹ or OR¹ group or together Z¹ andZ² link to form an optionally substituted 5- to 8-membered ring whichring is carbocyclic or heterocyclic, including one or more unsaturatedbonds, aryl or heteroaryl;Z² is H, an optionally substituted C₁-C₁₂ hydrocarbon group (preferablyan alkyl, alkenyl or aryl group), a 3-20 membered (preferably, a 5- to20-membered) heterocyclic group (preferably, a heteroaryl group) ortogether Z¹ and Z² are linked to form an optionally substituted 5- to8-membered ring (preferably, a 6- or 7-membered ring) which iscarbocyclic or heterocyclic, including aryl or heteroaryl;R¹ and R² are each independently absent, H, an optionally substitutedC₁-C₆ alkyl, alkene or alkyne group, an optionally substituted aryl orheterocyclic group (preferably, including a heteroaryl group) or NR¹R²is an optionally protected amino acid group where R¹ is H or a C₁-C₃alkyl group and R² is a group preferably obtained from a D- or L-aminoacid according to the chemical structure:

where R³ is a side chain derived from an amino acid preferably selectedfrom the group consisting of alanine (methyl), arginine(propyleneguanidine), asparagine (methylenecarboxyamide), aspartic acid(ethanoic acid), cysteine (thiol, reduced or oxidized di-thiol),glutamine (ethylcarboxyamide), glutamic acid (propanoic acid), glycine(H), histidine (methyleneimidazole), isoleucine (1-methylpropane),leucine (2-methylpropane), lysine (butyleneamine), ornithine(propyleneamine), methionine (ethylmethylthioether), phenylalanine(benzyl), proline (R³ forms a cyclic ring with R_(a) and the adjacentnitrogen group to form a pyrrolidine group), hydroxyproline, serine(methanol), threonine (ethanol, 1-hydroxyethane), tryptophan(methyleneindole), tyrosine (methylene phenol) or valine (isopropyl);X¹ is an optionally substituted S-alkyl (preferably, C₁-C₇ alkyl,preferably, S-Me or an amino acid group obtained from cysteine), anoptionally substituted S-aryl or an optionally substitutedS-heterocyclyl (preferably, S-heteroaryl), an optionally substitutedO-alkyl (preferably, C₁-C₇ alkyl, preferably. O-Me or an amino acidgroup obtained from serine), an optionally substituted O-aryl (or anoptionally substituted O-heterocyclyl (preferably, S-heteroaryl), aNR¹R² group where R¹ and R² are each independently H (often, R² is H),an optionally substituted alkyl group (preferably, a C₁-C₇ alkyl groupor an amino acid group obtained from lysine or ornithine), an optionallysubstituted aryl group or an optionally substituted heterocyclyl(preferably an optionally substituted heteroaryl), or (X is) an aminoacid group preferably obtained from a D- or L-amino acid according tothe chemical structure:

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline) and R³ is a side chain derived from an amino acidpreferably selected from the group consisting of alanine (methyl),arginine (propyleneguanidine), asparagine (methylenecarboxyamide),aspartic acid (ethanoic acid), cysteine (thiol, reduced or oxidizeddi-thiol), glutamine (ethylcarboxyamnide), glutamic acid (propanoicacid), glycine (H), histidine (methyleneimidazole), isoleucine(1-methylpropane), leucine (2-methylpropane), lysine (butyleneamine),methionine (ethylmethylthioether), phenylalanine (benzyl), proline (R³forms a cyclic ring with R_(a) and the adjacent nitrogen group to form apyrrolidine group), hydroxyproline, serine (methanol), threonine(ethanol, 1-hydroxyethane), tryptophan (methyleneindole), tyrosine(methylene phenol) or valine (isopropyl) (where the R³ side chain isoptionally protected). Preferably X is an optionally substitutedS—(C₁-C₁₂) alkyl (preferably, S-Me), O—(C₁-C₁₂) alkyl or a NH—R¹ groupwhere R¹ is an optionally substituted C₁-C₁₂, preferably a C₁-C₁₀ alkylgroup (preferably, NHR¹ is an ornithine or lysine moiety; andY′ is H or OH (preferably OH),or a salt form (especially including a pharmaceutically acceptable saltform), stereoisomer (including a diastereomer or enantiomer), solvate orpolymorph thereof.

In additional embodiments, the present invention is directed to apharmaceutical composition comprising an effective amount of a compoundas set forth above in combination with a pharmaceutically acceptablecarrier, additive and/or excipient, including an additional bioactiveagent, especially a bioactive agent useful in the treatment of adiabetic disorder or to inhibit or treat disorders related to the agingprocess as otherwise described herein.

In still further embodiments, the present invention is directed to amethod for treating a diabetic disorder or treating aging and/ordisorders related to the aging process in a patient in need comprisingadministering a pharmaceutical composition in an effective amount tosaid patient. Diabetic disorders which may be treated usingpharmaceutical compositions according to the present invention includetype I and type II diabetes, insulin resistance, glucose intolerance,insulin resistance syndrome, metabolic syndrome and related diseasestates and conditions including cardiovascular diseases associated withsame, hypertension, atherosclerosis, congestive heart failure, stroke,gallbladder disease, osteoarthritis, sleep apnea, reproductive disorderssuch as polycystic ovarian syndrome, cancers of the breast, prostate,and colon, and increased incidence of complications of generalanesthesia, as well as disorders such as infections, varicose veins,acanthosis nigricans, eczema, exercise intolerance,hypercholesterolemia, cholelithiasis, orthopedic injury andthromboembolic disease.

Additional embodiments of the present invention may be readily gleanedfrom a review of the detailed description of the present invention, setforth herein below.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows glucosepane and the iso-imidazole. (A) Chemical structureof the protein-bound glucosepane crosslink, depicting both non-aromaticiso-imdazole (1) and aromatic imidazole (2) tautomers. (B) Proposedbiosynthetic pathway for glucosepane crosslinks.

FIG. 2 shows a Retrosynthetic analysis of glucosepane to facilitatesynthesis.

FIG. 3 shows scheme 1 which shows the exemplary chemical synthetic stepswhich provide the protected ketone intermediate 14 from the protectedepoxide 8.

FIG. 4, scheme 2A, B and C show further analysis and chemical steps fromketone intermediate 14 to refine the chemical synthetic procedure of thepresent invention.

FIG. 5, scheme 3, shows the chemical steps to provide the fullyprotected glucosepane derivative 28 which can be deprotected aspresented to provide glucosepane derivative 5 as the formate or TFAsalt.

FIG. 6, scheme 3B, shows the synthesis of two additional2,4-diaminoimidazoles (29 and 30) from intermediates 26 and 27.

FIG. 7 shows 2D NOESY with t_(m)=1000 ms.

FIG. 8 shows a pH titration of glucosepane.

DETAILED DESCRIPTION OF THE INVENTION

In accordance with the present invention there may be employedconventional chemical synthetic and pharmaceutical formulation methods,as well as pharmacology, molecular biology, microbiology, andrecombinant DNA techniques within the skill of the art. Such techniquesare well-known and are otherwise explained fully in the literature.

Where a range of values is provided, it is understood that eachintervening value, to the tenth of the unit of the lower limit unlessthe context clearly dictates otherwise (such as in the case of a groupcontaining a number of carbon atoms), between the upper and lower limitof that range and any other stated or intervening value in that statedrange is encompassed within the invention. The upper and lower limits ofthese smaller ranges may independently be included in the smaller rangesis also encompassed within the invention, subject to any specificallyexcluded limit in the stated range. Where the stated range includes oneor both of the limits, ranges excluding either both of those includedlimits are also included in the invention.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. Although any methods andmaterials similar or equivalent to those described herein can also beused in the practice or testing of the present invention, the preferredmethods and materials are now described.

It is to be noted that as used herein and in the appended claims, thesingular forms “a,” “an”, “and” and “the” include plural referencesunless the context clearly dictates otherwise.

The following terms, among others, are used to describe the presentinvention. It is to be understood that a term which is not specificallydefined is to be given a meaning consistent with the use of that termwithin the context of the present invention as understood by those ofordinary skill.

The term “compound”, as used herein, unless otherwise indicated, refersto any specific chemical compound disclosed herein and includestautomers, regioisomers, geometric isomers, and where applicable,optical isomers (e.g. enantiomers), stereoisomers (e.g., diastereomers,such term also subsuming enantiomers and other diastereomers) thereof,as well as pharmaceutically acceptable salts and alternative salt formsas well as non-salt forms (depending upon the environment and/or pH inwhich the compound is found) and derivatives (including prodrug forms)thereof. Within its use in context, the term compound generally refersto a single compound, but also may include other compounds such asstereoisomers, regioisomers and/or optical isomers (including racemicmixtures) as well as specific enantiomers or enantiomerically enrichedmixtures of disclosed compounds as well as diastereomers and epimers,where applicable in context. The term also refers, in context to prodrugforms of compounds which have been modified to facilitate theadministration and delivery of compounds to a site of activity.

As used herein, the term “glucosepane” means a composition comprisingabout at least about 50%, at least about 55%, at least about 60%,preferably at least about 75%, at least about 80%, 85%, 90%, 95%, 98%,99% and 99-% pure glucosepane (see compound 1 of FIG. 1). Note that aglucosepane mixture may contain up to 8 enantiomeric and diastereomericforms of glucosepane. A glucosepane derivative is defined similarly withrespect to the relative amounts of its isomers (enantiomers and/ordiastereomers) in the final mixture of the composition. The term“stereoisomer” is used to refer to each of the up to 8 enantiomers anddiastereomeric forms of glucosepane, as well as other stereisomers ofglucosepane.

The term “patient” or “subject” is used throughout the specificationwithin context to describe an animal, generally a mammal and preferablya human, to whom treatment, including prophylactic treatment(prophylaxis), with the compositions according to the present inventionis provided. For treatment of those infections, conditions or diseasestates which are specific for a specific animal such as a human patient,the term patient refers to that specific animal.

The symbol

is used in chemical compounds according to the present invention tosignify that a bond between atoms is a single bond or double bondaccording to the context of the bond's use in the compound, whichdepends on the atoms (and substituents) used in defining the presentcompounds. Thus, where a carbon (or other) atom is used and the contextof the use of the atom calls for a double bond or single bond to linkthat atom with an adjacent atom in order to maintain the appropriatevalence of the atoms used, then that bond is considered a double bond ora single bond.

A diabetic disorder includes type I and type II diabetes, insulinresistance, glucose intolerance, insulin resistance syndrome, metabolicsyndrome and related disease states and conditions includingcardiovascular diseases associated with same, hypertension,atherosclerosis, congestive heart failure, stroke, gallbladder disease,osteoarthritis, sleep apnea, reproductive disorders such as polycysticovarian syndrome, cancers of the breast, prostate, and colon, andincreased incidence of complications of general anesthesia, as well asdisorders such as infections, varicose veins, acanthosis nigricans,eczema, exercise intolerance, hypercholesterolemia, cholelithiasis,orthopedic injury and thromboembolic disease.

The term “effective” is used herein, unless otherwise indicated, todescribe an amount of a compound or composition which, in context, isused to produce or effect an intended result, whether that resultrelates to the inhibition of the effects of diabetes or a diabeticcondition or disorder, to inhibit or reverse the effects of aging, or topotentiate the effects of a supplementary treatment used in treatingdiabetes or a diabetic condition or disorder or the effects of aging.This term subsumes all other effective amount or effective concentrationterms (including the term “therapeutically effective”) which areotherwise described in the present application.

The terms “treat”, “treating”, and “treatment”, etc., as used herein,refer to any action providing a benefit to a patient at risk for orafflicted by a diabetic disorder or condition, including lessening orsuppression of at least one symptom of diabetes or a diabetic disorder,delay in progression of diabetes, a diabetic disorder or the effects ofaging. Treatment, as used herein within context, may encompass bothprophylactic and therapeutic treatment depending on the context of theuse of the term. Treatment is established through the use of the term“treating” or “inhibiting” whereas prophylaxis is usually establishedthrough the use of the term “reducing the likelihood of”.

The term “pharmaceutically acceptable salt” or “salt” is used throughoutthe specification to describe a salt form of one or more of thecompositions herein which are presented preferably to increase thesolubility of the compound in a solvent, for example, in saline forparenteral delivery or in the gastric juices of the patient'sgastrointestinal tract in order to promote dissolution and thebioavailability of the compounds. Pharmaceutically acceptable saltsinclude those derived from pharmaceutically acceptable inorganic ororganic bases and acids. Suitable salts include those derived fromalkali metals such as potassium and sodium, alkaline earth metals suchas calcium, magnesium and ammonium salts, among numerous other acidswell known in the pharmaceutical art. Sodium and potassium salts may bepreferred as neutralization salts of carboxylic acids and free acidphosphate containing compositions according to the present invention.The term “salt” shall mean any salt consistent with the use of thecompounds according to the present invention. In the case where thecompounds are used in pharmaceutical indications, the term “salt” shallmean a pharmaceutically acceptable salt, consistent with the use of thecompounds as pharmaceutical agents.

The term “co-administration” shall mean that at least two compounds orcompositions are administered to the patient at the same time, such thateffective amounts or concentrations of each of the two or more compoundsmay be found in the patient at a given point in time. Although compoundsaccording to the present invention may be co-administered to a patientat the same time, or at slightly different intervals, the term embracesboth administration of two or more agents at the same time or atdifferent times, including sequential administration. Preferably,effective concentrations of all co-administered compounds orcompositions are found in the subject at a given time.

For example, compounds according to the present invention may beadministered with one or more agents that are useful in treatingdiabetes or a diabetic disorder or have an effect on the process ofaging in a patient. The type of co-administered agent can vary widelydepending on the particular clinical context. For example,co-administered agents can include anti-coagulant or coagulationinhibitory agents, anti-platelet or platelet inhibitory agents, thrombininhibitors, thrombolytic or fibrinolytic agents, anti-arrhythmic agents,anti-hypertensive agents, calcium channel blockers (L-type and T-type),cardiac glycosides, diuretics, mineralocorticoid receptor antagonists,phosphodiesterase inhibitors, cholesterol/lipid lowering agents andlipid profile therapies, traditional anti-diabetic agents,anti-depressants, anti-inflammatory agents (steroidal andnon-steroidal), anti-osteoporosis agents, hormone replacement therapies,oral contraceptives, anti-obesity agents, anti-anxiety agents,anti-proliferative agents, anti-tumor agents, anti-ulcer andgastroesophageal reflux disease agents, growth hormone and/or growthhormone secretagogues, thyroid mimetics (including thyroid receptorantagonist), anti-infective agents, anti-viral agents, anti-bacterialagents, anti-fungal agents and mixtures thereof.

In the case of Type II diabetes, useful additional agents include butare not limited to biguanides (e.g., metformin), glucosidase inhibitors(e.g., acarbose), insulins (including insulin secretagogues or insulinsensitizers), meglitinides (e.g., repaglinide), sulfonylureas (e.g.,glimepiride, glyburide and glipizide), biguanide/glyburide combinations(e.g., glucovance), thiozolidinediones (e.g., troglitazone,rosiglitazone and pioglitazone), PPAR-alpha agonists, PPAR-gammaagonists, PPAR alphaigamma dual agonists, SGLT2 inhibitors, inhibitorsof fatty acid binding protein (aP2), glucagon-like peptide-1 (GLP-1),and dipeptidyl peptidase IV (DP4) inhibitors and mixtures thereof.

The term “acyl” is art-recognized and refers to a group represented bythe general formula hydrocarbylC(O)—, preferably alkylC(O)—.

The term “acylamino” is art-recognized and refers to a moiety having anamino group and an acyl group and may include substitutents on same asotherwise disclosed herein. The term “alkylamino” refers to an aminogroup and an optionally substituted alkyl group on same as otherwisedescribed herein. “Dialkyl amino refers to an amino group and twooptionally substituted alkyl groups.

The term “aliphatic group” refers to a straight-chain, branched-chain,or cyclic aliphatic hydrocarbon group and includes saturated andunsaturated aliphatic groups, such as an alkyl group, an alkenyl group,and an alkynyl group. The term “alkenyl”, as used herein, refers to anoptionally substituted aliphatic group containing at least one doublebond, the substituted alkenyl moieties having substituents replacing ahydrogen on one or more carbons of the alkenyl group. Such substituentsmay occur on one or more carbons that are included or not included inone or more double bonds. Moreover, such substituents include all thosecontemplated for alkyl groups, as discussed herein, except wherestability of the moiety is prohibitive. For example, substitution ofalkenyl groups by one or more alkyl, carbocyclyl, aryl, heterocyclyl, orheteroaryl groups, among others is contemplated.

The terms “alkoxyl” or “alkoxy” as used herein refers to an alkyl group,as defined below, having an oxygen radical attached thereto.Representative alkoxyl groups include methoxy, ethoxy, propyloxy,tert-butoxy and the like.

An “ether” is two hydrocarbons covalently linked by an oxygen.Accordingly, the substituent of an alkyl that renders that alkyl anether is or resembles an alkoxyl, such as can be represented by one of—O-alkyl, —O-alkenyl, —O-alkynyl, —O—(CH₂)_(m)-substituent, where m is 0to 10, preferably 1 to 6, and the substituent is an aryl or substitutedaryl group, a cycloalkyl group, a cycloalkenyl, a heterocycle or apolycycle (two or three ringed), each of which may be optionallysubstituted.

The term “alkyl” refers to the radical of saturated aliphatic groups,including straight-chain alkyl groups, branched-chain alkyl groups,cycloalkyl (alicyclic) groups, alkyl-substituted cycloalkyl groups, andcycloalkyl-substituted alkyl groups. In preferred embodiments, astraight chain or branched chain alkyl has 10 or fewer carbon atoms inits backbone (e.g., C₁-C₁₂ for straight chains, C₁-C₁₂ for branchedchains), and more preferably 8-10 or fewer (C₁-C₁₀), and most preferably6 or fewer (C₁-C₆). Likewise, preferred cycloalkyls have from 3-10carbon atoms in their ring structure, and more preferably have 5, 6, 7or 8 carbons in the ring structure.

Moreover, the term “alkyl” (or “lower alkyl”) as used throughout thespecification, examples, and claims is intended to include both“unsubstituted alkyls” and “substituted alkyls”, the latter of whichrefers to alkyl moieties having substituents replacing a hydrogen on oneor more carbons of the hydrocarbon backbone. Such substituents caninclude, for example, a halogen, a hydroxyl, alkoxy (C₁-C₈, preferablyC₁-C₆) a carbonyl C₁-C₈ (such as a carboxyl, an alkoxycarbonyl ester, anoxycarbonyl ester, a formyl, or an acyl), a thiocarbonyl (such as athioester, a thioacetate, or a thioformate), a phosphoryl, a phosphate,a phosphonate, a phosphinate, an amino, including an amino incombination with a carboxylic acid (e.g., forming an amino acidsidechain such as an ornithine or lysine sidechain), an amido, anamidine, an imine, a cyano, a nitro, an azido, a sulfhydryl, analkylthio, a sulfate, a sulfonate, a sulfamoyl, a sulfonamido, asulfonyl, a heterocyclyl, an aralkyl, or an aromatic or heteroaromaticmoiety or as otherwise described herein. It will be understood by thoseskilled in the art that the individual substituent chemical moieties canthemselves be substituted. For instance, the substituents of asubstituted alkyl may include substituted and unsubstituted forms ofamino, azido, imino, amido, phosphoryl (including phosphonate andphosphinate), sulfonyl (including sulfate, sulfonamido, sulfamoyl andsulfonate), and silyl groups, as well as ethers, alkylthios, carbonyls(including ketones, aldehydes, carboxylates, and esters), —CF₃, —CN,nitro and the like. Exemplary, non-limiting substituted alkyls aredescribed herein. Cycloalkyls can be further substituted with alkyls,alkenyls, alkynyls, alkoxys, alkylthios, aminoalkyls,carbonyl-substituted alkyls, —CF₃, —CN, nitro, alkoxy and the like.

Analogous substitutions can be made to alkenyl and alkynyl groups toproduce, for example, without limitation, aminoalkenyls, aminoalkynyls,amidoalkenyls, amidoalkynyls, iminoalkenyls, iminoalkynyls,thioalkenyls, thioalkynyls, carbonyl-substituted alkenyls or alkynyls.

Unless the number of carbons is otherwise specified, “lower alkyl” asused herein means an alkyl group, as defined above, but having from oneto eight carbons, more preferably from one to six carbon atoms in itsbackbone structure. Likewise, “lower alkenyl” and “lower alkynyl” havesimilar chain lengths. Throughout the application, preferred alkylgroups are lower alkyls. In preferred embodiments, a substituentdesignated herein as alkyl is a lower alkyl.

The term “alkynyl”, as used herein, refers to an aliphatic groupcontaining at least one triple bond and is intended to include both“unsubstituted alkynyls” and “substituted alkynyls”, the latter of whichrefers to alkynyl moieties having substituents replacing a hydrogen onone or more carbons of the alkynyl group. Such substituents may occur onone or more carbons that are included or not included in one or moretriple bonds. Moreover, such substituents include all those contemplatedfor alkyl groups, as discussed above, except where stability isprohibitive. For example, substitution of alkynyl groups by one or morealkyl, carbocyclyl, aryl, heterocyclyl, or heteroaryl groups iscontemplated.

The term “alkylthio” refers to an alkyl group, as defined above, havinga sulfur radical attached thereto. In preferred embodiments, the“alkylthio” moiety is represented by one of —S-alkyl, —S-alkenyl,—S-alkynyl, and —S—(CH₂)_(m)-substituent, wherein m is 0 or an integerfrom 1 to 12, preferably 1 to 8 and substituent is the same as definedherein and as otherwise below (R₉ and R₁₀ for amine/amino).Representative alkylthio groups include methylthio, ethylthio, and thelike.

The terms “amine” and “amino” are art-recognized and refer to bothunsubstituted and substituted amines, e.g., a moiety that can berepresented, without limitation, by the general formula:

wherein R₉, R₁₀ and R′₁₀ each independently represent a hydrogen, anoptionally substituted alkyl, an alkenyl, —(CH₂)_(m)—R₈, or R₉ and R₁₀taken together with the N atom to which they are attached complete aheterocycle having from 4 to 8 atoms in the ring structure; R₈represents an aryl, a cycloalkyl, a cycloalkenyl, a heterocycle,including a heteroaryl or a polycycle, and m is zero or an integer inthe range of 1 to 8. In preferred embodiments, only one of R₉ or R₁₀ canbe a carbonyl, e.g., R₉, R₁₀ and the nitrogen together do not form animide. In certain such embodiments, neither R₉ and R₁₀ is attached to Nby a carbonyl, e.g., the amine is not an amide or imide, and the amineis preferably basic, e.g., its conjugate acid has a pK_(a) above 7. Ineven more preferred embodiments, R₉ and R₁₀ (and optionally, R′₁₀) eachindependently represent a hydrogen, an alkyl, an alkenyl, or—(CH₂)_(m)—R₈. Thus, the term “alkylamine” as used herein means an aminegroup, as defined above, having a substituted or unsubstituted alkylattached thereto, i.e., at least one of R₉ and R₁₀ is an alkyl group.Each of the groups which is bonded to the amine group, where applicable,may be optionally substituted. In certain preferred aspects the aminetogether with its substituent forms a lysine or ornithine radical.

The term “amido” is art-recognized as an amino-substituted carbonyl andincludes a moiety that can be represented by the general formula:

wherein R₉, R₁₀ are as defined above. Preferred embodiments of the amidewill not include imides that may be unstable.

The term “aralkyl”, as used herein, refers to an alkyl group substitutedwith an aryl group (e.g., an aromatic or heteroaromatic group).

The term “aryl” as used herein inn context includes 5-, 6-, and7-membered single-ring or aromatic groups which contain from zero tofour heteroatoms depending on the context of the term use, for example,benzene, pyrrole, furan, thiophene, imidazole, oxazole, thiazole,triazole, pyrazole, pyridine, pyrazine, pyridazine and pyrimidine, andthe like. Those aryl groups having heteroatoms in the ring structure mayalso be referred to as “aryl heterocycles”, “heteroaromatics” or“heteroaryl groups”. The aromatic ring can be substituted at one or morering positions with such substituents as otherwise described herein, forexample, halogen, azide, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl,polycyclyl, hydroxyl, alkoxyl, amino, nitro, sulfhydryl, imino, amido,phosphate, phosphonate, phosphinate, carbonyl, carboxyl, silyl, ether,alkylthio, sulfonyl, sulfonamido, ketone, aldehyde, ester, heterocyclyl,aromatic or heteroaromatic moieties, —CF₃, —CN, or the like. The term“aryl” also includes polycyclic ring systems having two or more cyclicrings in which two or more carbons are common to two adjoining rings(the rings are “fused rings”) wherein at least one of the rings isaromatic, e.g., the other cyclic rings can be cycloalkyls,cycloalkenyls, cycloalkynyls, aryls and/or heterocyclyls.

The term “carbocycle”, as used herein, refers to an aromatic ornon-aromatic ring in which each atom of the ring is carbon.

The term “carbonyl” is art-recognized and includes such moieties as canbe represented by the general formula:

wherein X is a bond or represents an oxygen or a sulfur, and R₁₁represents, for example without limitation, a hydrogen, an alkyl, analkenyl, —(CH₂)_(m)—R₈ or a pharmaceutically acceptable salt, R′₁₁represents a hydrogen, an alkyl, an alkenyl or —(CH₂)_(m)—R₈, where m(0-8) and R₈ are as otherwise described herein without limitation. WhereX is oxygen and R₁₁ or R′₁₁ is not hydrogen, the formula represents an“ester”. Where X is oxygen, and R₁₁ is as defined above, the moiety isreferred to herein as a carboxyl group, and particularly when R₁₁ ishydrogen, the formula represents a “carboxylic acid”. Where X is oxygen,and R′₁₁ is hydrogen, the formula represents a “formate”. In general,where the oxygen atom of the above formula is replaced by sulfur, theformula represents a “thiocarbonyl” group. Where X is sulfur and R₁₁ orR′₁₁ is not hydrogen, the formula represents a “thioester.” Where X issulfur and R₁₁ is hydrogen, the formula represents a “thiocarboxylicacid.” Where X is sulfur and R′₁₁ is hydrogen, the formula represents a“thiolformate.” On the other hand, where X is a bond, and R₁₁ is nothydrogen, the above formula represents a “ketone” group. Where X is abond, and R₁₁ is hydrogen, the above formula represents an “aldehyde”group.

The term “electron withdrawing group” refers to chemical groups whichwithdraw electron density from the atom or group of atoms to whichelectron withdrawing group is attached. The withdrawal of electrondensity includes withdrawal both by inductive and bydelocalization/resonance effects. Examples of electron withdrawinggroups attached to aromatic rings include perhaloalkyl groups, such astrifluoromethyl, halogens, azides, carbonyl containing groups such asacyl groups, cyano groups, and imine containing groups. The term“electron contributing group” refers to chemical groups which donateelectron density to the atom or group of atoms to which the electronicdonating group is attached. Examples of such groups include the alkoxyand amine groups, among others.

The term “ester”, as used herein, refers to a group —C(O)O-substituentwherein the substituent represents, for example, a hydrocarbyl or othersubstitutent as is otherwise described herein.

The terms “halo” and “halogen” as used herein means halogen and includeschloro, fluoro, bromo, and iodo.

The terms “hetaroalkyl” and “heteroaralkyl”, as used herein, refers toan alkyl group substituted with a hetaryl group.

The terms “heterocycle” or “heterocyclic group” refer to 3- to10-membered ring structures, more preferably 3- to 7-membered rings,whose ring structures include one to four heteroatoms. Heterocycles canalso be polycycles and can include up to 20 member polycyclic ringsystems. Heterocyclyl groups include, for example, without limitation,thiophene, thianthrene, furan, pyran, isobenzofuran, chromene, xanthene,phenoxathiin, pyrrole, imidazole, pyrazole, isothiazole, isoxazole,pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole,indole, indazole, purine, quinolizine, isoquinoline, quinoline,phthalazine, naphthyridine, quinoxaline, quinazoline, cinnoline,pteridine, carbazole, carboline, phenanthridine, acridine, pyrimidine,phenanthroline, phenazine, phenarsazine, phenothiazine, furazan,phenoxazine, pyrrolidine, oxolane, thiolane, oxazole, piperidine,piperazine, morpholine, lactones, lactams such as azetidinones andpyrrolidinones, sultams, sultones, and the like. The heterocyclic ringcan be substituted at one or more positions with such substituents asdescribed above without limitation, as for example, halogen, alkyl,aralkyl, alkenyl, alkynyl, cycloalkyl, hydroxyl, amino, nitro,sulthydryl, imino, amido, phosphate, phosphonate, phosphinate, carbonyl,carboxyl, silyl, ether, alkylthio, sulfonyl, ketone, aldehyde, ester, aheterocyclyl, an aromatic or heteroaromatic moiety, —CF₃, —CN, or thelike, and as otherwise described herein.

The terms “heteroaryl” and “hetaryl” include substituted orunsubstituted aromatic single ring structures, preferably 5- to7-membered rings, more preferably 5- to 6-membered rings, whose ringstructures include at least one heteroatom, preferably one to fourheteroatoms, more preferably one or two heteroatoms. The terms“heteroaryl” and “hetaryl” also may include up to 20-membered polycyclicring systems having two or more cyclic rings in which two or morecarbons are common to two adjoining rings wherein at least one of therings is heteroaromatic, e.g., the other cyclic rings can becycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/orheterocyclyls. Heteroaryl groups include, for example, pyrrole, furan,thiophene, imidazole, oxazole, thiazole, pyrazole, pyridine, pyrazine,pyridazine, and pyrimidine, and the like.

The term “heteroatom” as used herein means an atom of any element otherthan carbon or hydrogen. Preferred heteroatoms are nitrogen, oxygen, andsulfur.

Thus, the terms “heterocyclyl”, “heterocycle”, and “heterocyclic” referto substituted or unsubstituted aromatic or non-aromatic ring structures(which can be cyclic, bicyclic or a fused ring system), preferably 3- to10-membered rings, more preferably 3- to 7-membered rings, whose ringstructures include at least one heteroatom, preferably one to fourheteroatoms, more preferably one or two heteroatoms. The terms“heterocyclyl” and “heterocyclic” also include polycyclic ring systemshaving two or more cyclic rings in which two or more carbons are commonto two adjoining rings wherein at least one of the rings isheterocyclic, e.g., the other cyclic rings can be cycloalkyls,cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls.Heterocyclyl groups include, for example, piperidine, piperazine,pyrrolidine, morpholine, lactones, lactams, and the like.

The term “5- to 20-membered heterocyclic group” or “5- to 14-memberedheterocyclic group” as used throughout the present specification refersto an aromatic or non-aromatic cyclic group having 5 to 20 atoms,preferably 5 to 14 atoms forming the cyclic ring(s) and including atleast one hetero atom such as nitrogen, sulfur or oxygen among the atomsforming the cyclic ring, which is a “5 to 20-membered, preferably 5- to14-membered aromatic heterocyclic group” (also, “heteroaryl” or“heteroaromatic”) in the former case and a “5 to 20-membered”,preferably a “5- to 14-membered non-aromatic heterocyclic group” in thelatter case.

Among the heterocyclic groups which may be mentioned includenitrogen-containing aromatic heterocycles such as pyrrole, pyridine,pyridone, pyridazine, pyrimidine, pyrazine, pyrazole, imidazole,triazole, tetrazole, indole, isoindole, indolizine, purine, indazole,quinoline, isoquinoline, quinolizine, phthalazine, naphthyridine,quinoxaline, quinazoline, cinnoline, pteridine, imidazopyridine,imidazotriazine, pyrazinopyridazine, acridine, phenanthridine,carbazole, carbazoline, perimidine, phenanthroline, phenacene,oxadiazole, benzimidazole, pyrrolopyridine, pyrrolopyrimidine andpyridopyrimidine: sulfur-containing aromatic heterocycles such asthiophene and benzothiophene; oxygen-containing aromatic heterocyclessuch as furan, pyran, cyclopentapyran, benzofuran and isobenzofuran; andaromatic heterocycles comprising 2 or more hetero atoms selected fromamong nitrogen, sulfur and oxygen, such as thiazole, thiadizole,isothiazole, benzoxazole, benzothiazole, benzothiadiazole,phenothiazine, isoxazole, furazan, phenoxazine, pyrazoloxazole,imidazothiazole, thienofuran, furopyrrole, pyridoxazine, furopyridine,furopyrimidine, thienopyrimidine and oxazole.

As examples of the “5- to 14-membered aromatic heterocyclic group” theremay be mentioned preferably, pyridine, triazine, pyridone, pyrimidine,imidazole, indole, quinoline, isoquinoline, quinolizine, phthalazine,naphthyridine, quinazoline, cinnoline, acridine, phenacene, thiophene,benzothiophene, furan, pyran, benzofuran, thiazole, benzthiazole,phenothiazine, pyrrolopyrimidine, furopyridine and thienopyrimidine,more preferably pyridine, thiophene, benzothiophene, thiazole,benzothiazole, quinoline, quinazoline, cinnoline, pyrrolopyrimidine,pyrimidine, furopyridine and thienopyrimidine. The term “heterocyclicgroup” shall generally refer to 3 to 20-membered heterocyclic groups,preferably 3 to 14-membered heterocyclic groups and all subsets ofheterocyclic groups (including non-heteroaromatic or heteroaromatic)subsumed under the definition of heterocyclic group are 3 to 20-memberedheterocyclic groups, preferably 3 to 14-membered heterocyclic groups.

The term “8 to 20-membered heterocyclic group”, or “8 to 14-memberedheterocyclic group” refers to an aromatic or non-aromatic fused bicyclicor tricyclic group having 8 to 20, preferably 8 to 14 atoms forming thecyclic rings (two or three rings) and include at least one hetero atomsuch as nitrogen, sulfur or oxygen among the atoms forming the cyclicrings, which is a “8 to 20-membered”, preferably a “8- to 14-memberedaromatic heterocyclic group” (also, “heteroaryl” or “heteroaromatic”) inthe former case and a “8 to 20-membered”, preferably a “8- to14-membered non-aromatic heterocyclic group” in the latter case. “8 to20-membered heterocyclic groups” and “8 to 14 membered heterocyclicgroups” are represented by fused bicyclic, tricyclic and tetracyclicring structures containing nitrogen atoms such as indole, isoindole,indolizine, purine, indazole, quinoline, isoquinoline, quinolizine,phthalazine, naphthyridine, quinoxaline, quinazoline, cinnoline,pteridine, imidazopyridine, imidazotriazine, pyrazinopyridazine,acridine, phenanthridine, carbazole, carbazoline, perimidine,phenanthroline, phenacene, benzimidazole, pyrrolopyridine,pyrrolopyrimidine and pyridopyrimidine; sulfur-containing aromaticheterocycles such as thiophene and benzothiophene; oxygen-containingaromatic heterocycles such as cyclopentapyran, benzofuran andisobenzofuran; and aromatic heterocycles comprising 2 or more heteroatoms selected from among nitrogen, sulfur and oxygen, such asbenzoxazole, benzothiazole, benzothiadiazole, phenothiazine,benzofurazan, phenoxazine, pyrazoloxazole, imidazothiazole, thienofuran,furopyrrole, pyridoxazine, furopyridine, furopyrimidine andthienopyrimidine, among others.

The term “5- to 14-membered non-aromatic heterocyclic group” as usedthroughout the present specification refers to non-aromatic cyclic grouphaving 5 to 14 atoms forming the cyclic ring and including at least onehetero atom such as nitrogen, sulfur or oxygen among the atoms formingthe cyclic ring. As specific examples there may be mentionednon-aromatic heterocycles such as pyrrolidinyl, pyrrolinyl, piperidinyl,piperazinyl. N-methylpiperazinyl, imidazolinyl, pyrazolidinyl,imidazolidinyl, morpholinyl, tetrahydropyranyl, azetidinyl, oxetanyl,oxathiolanyl, pyridone, 2-pyrrolidone, ethyleneurea, 1,3-dioxolane,1,3-dioxane, 1,4-dioxane, phthalimideandsuccinimide. As examples of the“5- to 14-membered non-aromatic heterocyclic group” there may bementioned preferably, pyrrolidinyl, piperidinyl and morpholinyl, andmore preferably pyrrolidinyl, piperidinyl, morpholinyl and pyrrole.

The term “8- to 14-membered non-aromatic heterocyclic group” as usedthroughout the present specification refers to a non-aromatic fusedcyclic ring system (generally with two or three rings) having 8 to 14atoms forming the cyclic rings (bicyclic or tricyclic) and including atleast one hetero atom such as nitrogen, sulfur or oxygen among the atomsforming the cyclic rings.

The term “5- to 14-membered heterocyclic group” as used throughout thepresent specification refers to an aromatic or non-aromatic cyclic grouphaving 5 to 14 atoms forming the cyclic ring and including at least onehetero atom such as nitrogen, sulfur or oxygen among the atoms formingthe cyclic ring, which is a “5- to 14-membered aromatic heterocyclicgroup” in the former case and a “5- to 14-membered non-aromaticheterocyclic group” in the latter case. Specific examples of the “5- to14-membered heterocyclic group” therefore include specific examples ofthe “5- to 14-membered aromatic heterocyclic group” and specificexamples of the “5- to 14-membered non-aromatic heterocyclic group”.

As the “5- to 14-membered heterocyclic group” there may be mentionedpreferably pyrrolidinyl, piperidinyl, morpholinyl, pyrrole, pyridine,pyridone, pyrimidine, imidazole, indole, quinoline, isoquinoline,quinolizine, phthalazine, naphthyridine, quinazoline, cinnoline,acridine, phenacene, thiophene, benzothiophene, furan, pyran,benzofuran, thiazole, benzothiazole, phenothiazine and carbostyryl, morepreferably pyrrolidinyl, piperidinyl, morpholinyl, pyrrole, pyridine,thiophene, benzothiophene, thiazole, benzothiazole, quinoline,quinazoline, cinnoline and carbostyryl, and even more preferablythiazole, quinoline, quinazoline, cinnoline and carbostyryl, amongothers.

The term “6- to 14-membered aromatic heterocyclic group” as usedthroughout the present specification refers to those substituentsdefined by “5- to 14-membered aromatic heterocyclic group” which have 6to 14 atoms forming the cyclic ring. As specific examples there may bementioned pyridine, pyridone, pyrimidine, indole, quinoline,isoquinoline, quinolizine, phthalazine, naphthyridine, quinazoline,cinnoline, acridine, benzothiophene, benzofuran, thiazole, benzothiazoleand phenothiazine*. “8 to 14-membered aromatic heterocyclic groups”refer to those substituents or radicals having 8 to 14 atoms formingfused two or three cyclic ring systems. Specific examples includeindole, quinoline, isoquinoline, quinolizine, phthalazine,naphthyridine, quinazoline, cinnoline, acridine, benzothiophene,benzofuran, benzothiazole, pyrrolopyrimidine, pyrrolopyrazine,furopyrimidine and phenothiazine, among numerous others.

The term “6- to 14-membered heterocyclic group” as used throughout thepresent specification refers to those substituents defined by “5- to14-membered heterocyclic group” which have 6 to 14 atoms forming thecyclic ring(s). As specific examples there may be mentioned piperidinyl,piperazinyl, N-methylpiperazinyl, morpholinyl, tetrahydropyranyl,1,4-dioxane and phthalimide.

The term “3 to 7-membered heterocyclic group” as used throughout thepresent specification refers to those heterocyclic substituents whichhave 3 to 7 atoms forming the cyclic ring, preferably 5 to 6 atomsforming the cyclic ring.

The term “8 to 14-membered heterocyclic group” as used throughout thepresent specification refers to those substituents defined “8- to14-membered heterocyclic groups which have 8 to 14 atoms forming thefused cyclic ring system.

The term “heterocyclylalkyl”, as used herein, refers to an alkyl groupsubstituted with a heterocycle group.

The term “hydrocarbyl”, as used herein, refers to an optionallysubstituted group that is bonded through a carbon atom and typically hasat least one carbon-hydrogen bond and a primarily carbon backbone, butmay optionally include heteroatoms. Hydrocarbyl groups include, but arenot limited to aryl, heteroaryl, carbocycle, heterocycle, alkyl,alkenyl, alkynyl, and combinations thereof.

The term “lower” when used in conjunction with a chemical moiety, suchas, acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy is meant toinclude groups where there are ten or fewer atoms in the substituent,preferably eight or fewer, more preferably six or fewer. A “loweralkyl”, for example, refers to an alkyl group that contains ten or fewercarbon atoms, preferably six or fewer. In certain embodiments, acyl,acyloxy, alkyl, alkenyl, alkynyl, or alkoxy substituents defined hereinare respectively lower acyl, lower acyloxy, lower alkyl, lower alkenyl,lower alkynyl, or lower alkoxy, whether they appear alone or incombination with other substituents, such as in the recitationshydroxyalkyl and aralkyl (in which case, for example, the atoms withinthe aryl group are not counted when counting the carbon atoms in thealkyl substituent).

As used herein, the term “nitro” means —NO₂; the term “halogen”designates —F, —Cl, —Br or —I; the term “sulfhydryl” means —SH: the term“hydroxyl” means —OH; and the term “sulfonyl” means —SO₂—.

The terms “polycyclyl” or “polycyclic group” refer to two or more rings(e.g., cycloalkyls, cycloalkenyls, cycloalkynyls, aryls and/orheterocyclyls) in which two or more atoms are common to two adjoiningrings, e.g., the rings are “fused rings”. Rings that are joined throughnon-adjacent atoms are termed “bridged” rings. Each of the rings of thepolycycle can be substituted with, without limitation, such substituentsas described above, as for example, halogen, alkyl, aralkyl, alkenyl,alkynyl, cycloalkyl, hydroxyl, amino, nitro, sulfhydryl, imino, amido,phosphate, phosphonate, phosphinate, carbonyl, carboxyl, silyl, ether,alkylthio, sulfonyl, ketone, aldehyde, ester, a heterocyclyl, anaromatic or heteroaromatic moiety, —CF3, —CN, or the like.

The phrase “protecting group” or “blocking group” as used herein meanstemporary substituents which protect a potentially reactive functionalgroup from undesired chemical transformations. Examples of suchprotecting groups include esters of carboxylic acids, silyl ethers ofalcohols, and acetals and ketals of aldehydes and ketones, respectively.The field of protecting group chemistry has been reviewed (Greene, T.W.; Wuts, P. G. M. Protective Groups in Organic Synthesis, 2^(nd) ed.;Wiley: New York, 1991). Typical blocking groups are used on alcoholgroups, amine groups, carbonyl groups, carboxylic acid groups, phosphategroups and alkyne groups among others.

Exemplary alcohol/hydroxyl protecting groups include acetyl (removed byacid or base), benzoyl (removed by acid or base), benzyl (removed byhydrogenolysis, β-methoxyethoxymethyl ether (MEM, removed by acid),dimethoxytrityl [bis-(4-methoxyphenyl)phenylmethyl] (DMT, removed byweak acid), methoxymethyl ether (MOM, removed by acid), methoxytrityl[(4-methoxyphenyl)diphenylmethyl], (MMT, Removed by acid andhydrogenolysis), p-methoxylbenzyl ether (PMB, removed by acid,hydrogenolysis, or oxidation), methylthiomethyl ether (removed by acid),pivaloyl (Piv, removed by acid, base or reductant agents. More stablethan other acyl protecting groups, tetrahydropyranyl (THP, removed byacid), tetrahydrofuran (THF, removed by acid), trityl (triphenyl methyl,(Tr, removed by acid), silyl ether (e.g. trimethylsilyl or TMS,tert-butyldimethylsilyl or TBDMS, tri-iso-propylsilyloxymethyl or TOM,and triisopropylsilyl or TIPS, all removed by acid or fluoride ion suchas such as NaF, TBAF (tetra-n-butylammonium fluoride, HF-Py, orHF-NEt₃): methyl ethers (removed by TMSI in DCM, MeCN or chloroform orby BBr₃ in DCM) or ethoxyethlyl ethers (removed by strong acid).

Exemplary amine-protecting groups include carbobenzyloxy (Cbz group,removed by hydrogenolysis), p-Methoxylbenzyl carbon (Moz or MeOZ group,removed by hydrogenolysis), tert-butyloxycarbonyl (BOC group, removed byconcentrated strong acid or by heating at elevated temperatures),9-Fluorenylmethyloxycarbonyl (FMOC group, removed by weak base, such aspiperidine or pyridine), acyl group (acetyl, benzoyl, pivaloyl, bytreatment with base), benzyl (Bn groups, removed by hydrogenolysis),carbamate, removed by acid and mild heating, p-methoxybenzyl (PMB,removed by hydrogenolysis), 3,4-dimethoxybenzyl (DMPM, removed byhydrogenolysis), p-methoxyphenyl (PMP group, removed by ammonium ceriumIV nitrate or CAN); tosyl (Ts group removed by concentrated acid andreducing agents, other sulfonamides, Mesyl. Nosyl & Nps groups, removedby samarium iodide, tributyl tin hydride.

Exemplary carbonyl protecting groups include acyclical and cyclicalacetals and ketals (removed by acid), acylals (removed by Lewis acids)and dithianes (removed by metal salts or oxidizing agents).

Exemplary carboxylic acid protecting groups include methyl esters(removed by acid or base), benzyl esters (removed by hydrogenolysis),tert-butyl esters (removed by acid, base and reductants), esters of2,6-disubstituted phenols (e.g. 2,6-dimethylphenol,2,6-diisopropylphenol, 2,6-di-tert-butylphenol, removed at roomtemperature by DBU-catalyzed methanolysis under high-pressureconditions, silyl esters (removed by acid, base and organometallicreagents), orthoesters (removed by mild aqueous acid), oxazoline(removed by strong hot acid (pH <1, T >100° C.) or strong hot alkali(pH >12, T >100° C.)).

Exemplary phosphate group protecting groups including cyanoethyl(removed by weak base) and methyl (removed by strong nucleophiles, e.g.thiophenol/TEA).

Exemplary terminal alkyne protecting groups include propargyl alcoholsand silyl groups.

The term “substituted” refers to moieties having substituents replacinga hydrogen on one or more carbons of the backbone. It will be understoodthat “substitution” or “substituted with” includes the implicit provisothat such substitution is in accordance with permitted valence of thesubstituted atom and the substituent, and that the substitution resultsin a stable compound, e.g., which does not spontaneously undergotransformation such as by rearrangement, cyclization, elimination, etc.

As used herein, the term “substituted” is contemplated to include allpermissible substituents of organic compounds. In a broad aspect, thepermissible substituents include acyclic and cyclic, branched andunbranched, carbocyclic and heterocyclic, aromatic, non-aromatic andinorganic substituents of organic compounds. The permissiblesubstituents can be one or more and the same or different forappropriate organic compounds. For purposes of this invention, theheteroatoms such as nitrogen may have hydrogen substituents and/or anypermissible substituents of organic compounds described herein whichsatisfy the valences of the heteroatoms. Substituents can include anysubstituents (groups) as otherwise described herein, for example, ahalogen, a hydroxyl, a carbonyl (such as a carboxyl, an alkoxycarbonyl,a formyl, or an acyl), an ether, a thioether, a thiocarbonyl (such as athioester, a thioacetate, or a thioformate), an alkoxyl, a phosphoryl, aphosphate, a phosphonate, a phosphinate, an amino, an amido, an amidine,an imine, a cyano, a nitro, an azido, a sulfhydryl, an alkylthio, asulfate, a sulfonate, a sulfamoyl, a sulfonamido, a sulfonyl, aheterocyclyl, an aralkyl, or an aromatic or heteroaromatic moiety. Itwill be understood by those skilled in the art that the moietiessubstituted on a moiety or chemical group can themselves be substituted.

It will be understood that “substitution” or “substituted with” includesthe implicit proviso that such substitution is in accordance withpermitted valence of the substituted atom and the substituent, and thatthe substitution results in a stable compound, e.g., which does notspontaneously undergo transformation such as by rearrangement,cyclization, elimination, etc. It is acknowledged that the term“unsubstituted” simply refers to a hydrogen substituent or nosubstituent within the context of the use of the term.

Preferred substituents for use in the present invention include, forexample, within context, hydroxyl, carboxyl, cyano (C≡N), nitro (NO₂),halogen (preferably, 1, 2 or 3 halogens, especially on an alkyl,especially a methyl group such as a trifluoromethyl), thiol, alkyl group(preferably, C₁-C₆, more preferably, C₁-C₃), alkoxy group (preferably,C₁-C₆ alkyl or aryl, including phenyl), ether (preferably, C₁-C₆ alkylor aryl), ester (preferably, C₁-C₆ alkyl or aryl) including alkyleneester (such that attachment is on the alkylene group, rather than at theester function which is preferably substituted with a C₁-C₆ alkyl oraryl group), thioether (preferably, C₁-C₆ alkyl or aryl) (preferably,C₁-C₆ alkyl or aryl), thioester (preferably, C₁-C₆ alkyl or aryl),halogen (F, Cl, Br, I), nitro or amine (including a five- orsix-membered cyclic alkylene amine, including a C₁-C₆ alkyl amine orC₁-C₆ dialkyl amine), alkanol (preferably, C₁-C₆ alkyl or aryl), oralkanoic acid (preferably, C₁-C₆ alkyl or aryl). More preferably, theterm “substituted” shall mean within its context of use alkyl, alkoxy,halogen, hydroxyl, carboxylic acid, nitro and amine (including mono- ordi-alkyl substituted amines). Any substitutable position in a compoundaccording to the present invention may be substituted in the presentinvention, but preferably no more than 5, more preferably no more than 3substituents are present on a single ring or ring system. Preferably,the term “unsubstituted” shall mean substituted with one or more Hatoms.

As used herein, the definition of each expression of alkyl, m, n, etc.when it occurs more than once in any structure, is intended to reflectthe independence of the definition of the same expression in thestructure.

By way of example, certain preferred aromatic and aliphatic rings andtheir derivatives and substituents which may be used as pharmacophoresor substituents in compounds according to the present invention include,but are not limited to, phenyl, benzyl, pyridine, cyclohexadiene,dihydropyridine, tetrahydropyridine, piperidine, pyrazine,tetrahydro-pyrazine, dihydro-pyrazine, piperazine, pyrimidine,dihydro-pyrimidine tetrahydro-pyrimidine, hexahydro-pyrimidine,pyrimidinone, triazine, dihydro-triazine, tetrahydro-triazine,triazinane, tetrazine, dihydro-tetrazine, tetrahydro-tetrazine,tetrazinane, pyrrol, dihydro-pyrrole, pyrrolidine, imidazolidine,dihydro-imidazolidine, imidazole, dihydro-imidazole, azetidine,triazole, dihydro-triazole, triazolidine, tetrazole, dihydro-tetrazole,tetrazolidine, diazepane, tetrahydro-diazepine, dihydro-diazepine,diazepine, oxazole, dihydrooxazole, oxazolidine, isoxazole,dihydroisoxazole, isoxazolidine, thiazole, dihydrothiazole,thiazolidine, isothiazole, dihydroisothiazole, isothiazolidine,oxadiazole, dihydro-oxadiazole, oxadiazolidine, thiadiazole,dihydro-thidiazole, thidiazolidine, oxazinane, dihydro-oxazinane,dihydro-oxazine, oxazine (including morpholine), thiazinane,dihydro-thiazinane, dihydro-thiazine, thiazine (includingthiomorpholine), thiazine, furan, dihydrofuran, tetrahydrofuran,thiophene, pyridazine-3,6-dione, tetrahydrothiophene, dihydrothiophene,tetrahydrothiophene, dithiolane, dithiole, dithiolone, dioxolane,dioxole, oxathiole, oxathiolane, pyridinone, dioxane, dioxanedione,benzoquinone, dihydro-dioxine, dioxine, pyran, 3,4-dihydro-2H-pyran,pyranone, 2H-pyran-2,3(4H)-dione, oxathiane, dihydro-oxathiine,oxathiine, oxetane, thietane, thiazeto, cyclohexadienone, lactam,lactone, piperazinone, pyrroledione, cyclopentenone, oxazete,oxazinanone, dioxolane, 3,4-dihydro-2H-thiopyran 1,1-dioxide,dioxolanone, oxazolidinone, oxazolone, thiane 1-oxide, thiazinane1-oxide, tetrahydro-thiopyran, thiane 1,1-dioxide, dioxazinane,pyrazolone, 1,3-thiazete, thiazinane 1,1-dioxide,6,7-dihydro-5H-1,4-dioxepine, 1,2-dihydropyridazin-3(4H)-one,pyridine-2,6(1H,3H)-dione, and sugar (glucose, mannose, galactose,fucose, fructose, ribose).

Bicyclic and fused rings include, for example, naphthyl, quinone,quinolinone, dihydroquinoline, tetrahydroquinoline, naphthyridine,quinazoline, dihydroquinazoline, tetrahydroquinazoline, quinoxaline,dihydroquinazoline, tetrahydroquinazoline, pyrazine,quinazoline-2,4(1H,3H)-dione, isoindoline-1,3-dione,octahydro-pyrrolo-pyridine, indoline, isoindoline, hexahydro-indolone,tetrahydropyrrolo oxazolone, hexahydro-2H-pyrrolo[3,4-d]isoxazole,tetrahydro-1,6-naphthyridine,2,3,4,5,6,7-hexahydro-1H-pyrrolo[3,4-c]pyridine, 1H-benzo[d]imidazole,octahydropyrrolo[3,4-c]pyrrole, 3-azabicyclo[3.1.0]hexane,7-azabicyclo[2.2.1]hept-2-ene, diazabicyclo-heptane, benzoxazole,indole, 1,4-diazabicyclo[3.3.1]nonane, azabicyclo-octane,naphthalene-1,4-dione, indene, dihydroindene,2,3,3a,7a-tetrahydro-1H-isoindole, 2,3,3a,4,7,7a-hexahydro-1H-isoindole,1,3-dihydroisobenzofuran, 1-methyl-3a,4,5,6,7,7a-hexahydro-1H-indole,3-azabicyclo[4.2.0]octane, 5,6-dihydrobenzo[b]thiophene,5,6-dihydro-4H-thieno[2,3-b]thiopyran, 3,4-dihydropyrazin-2(1H)-one,2H-benzo[b][1,4]thiazine, naphthyridin-4(1H)-one,octahydropyrrolo[1,2-a]pyrazine, imidazo-pyridazine,tetrahydroimidazo-pyridazine, tetrahydropyridazine, thiazinone,5-thia-1-azabicyclo[4.2.0]oct-2-en-8-one,4-thia-1-azabicyclo[3.2.0]heptan-7-one,1,6,7,8-tetrahydroimidazo[4,5-d][1,3]diazepine,8H-thiazolo[4,3-c][1,4]oxazin-4-ium,8H-thiazolo[4,3-c][1,4]thiazin-4-ium, pteridine,thiazolo[3,4-a]pyrazin-4-ium,7-(methylimino)-7H-pyrrolo[1,2-c]thiazol-4-ium, thiazolo-pyrazine,3,7-dioxabicyclo[4.1.0]hept-4-ene,6,7-dihydro-5H-pyrrolo[1,2-a]imidazole,3,3a-dihydrofuro[3,2-b]furan-2(6aH)-one,tetrahydro-3aH-[1,3]dioxolo[4,5-c]pyrrole,7-ethylidene-7H-pyrrolo[1,2-c]thiazol-4-ium,hexahydro-1H-pyrrolo[2,1-c][1,4]oxazine,6,7,8,8a-tetrahydro-1H-pyrrolo[2,1-c][1,4]oxazine,2-azabicyclo[2.2.2]oct-2-ene, 6,6a-dihydrothieno[3,2-b]furan-5(3aH)-one,4,5-dihydropyridin-3(2H)-one, 4,7a-dihydro-3aH-[1,3]dioxolo[4,5-c]pyran,6,7-dihydro-1H-furo[3,4-c]pyran-1,3(4H)-dione,3,3a,4,7a-tetrahydro-2H-furo[2,3-b]pyran,2,4a,7,7a-tetrahydro-1H-cyclopenta[c]pyridine,4H-pyrano[3,2-b]pyridine-4,8(5H)-dione,1,2,3,3a,4,7a-hexahydropyrano[4,3-b]pyrrole,2,3,8,8a-tetrahydroindolizin-7(1H)-one,octahydro-1H-pyrido[1,2-a]pyrazin-1-one,2,6,7,8,9,9a-hexahydro-1H-pyrido[1,2-a]pyrazin-1-one,6,7,8,8a-tetrahydropyrrolo[1,2-a]pyrazin-1(2H)-one,hexahydropyrrolo[1,2-a]pyrazin-1(2H)-one, bicyclo[2.2.1]hepta-2,5-diene.

Spiro moieties: 1,5-dioxaspiro[5.5]undecane, 1,4-dioxaspiro[4.5]decane,1,4-diazabicyclo[3.2.1]octane, 5-azaspiro[2.5]octane,5-azaspiro[2.4]heptane, 3,9-diaza-6-azoniaspiro[5.5]undecane,3,4-dihydrospiro[benzo[b][1,4]oxazine-2,1′-cyclohexane],7-oxa-4-azaspiro[2.5]oct-5-ene.

Pharmaceutical compositions comprising combinations of an effectiveamount of at least one glucosepane or glucosepane derivative compound oran imidazole compound according to the present invention otherwisedescribed herein, all in effective amounts, in combination with apharmaceutically effective amount of a carrier, additive or excipient,represents a further aspect of the present invention. Optionally, atleast one additional bioactive agent may be included in pharmaceuticalcompositions according to the present invention.

The compositions used in methods of treatment of the present invention,and pharmaceutical compositions of the invention, may be formulated in aconventional manner using one or more pharmaceutically acceptablecarriers and may also be administered in controlled-releaseformulations. Pharmaceutically acceptable carriers that may be used inthese pharmaceutical compositions include, but are not limited to, ionexchangers, alumina, aluminum stearate, lecithin, serum proteins, suchas human serum albumin, buffer substances such as phosphates, glycine,sorbic acid, potassium sorbate, partial glyceride mixtures of saturatedvegetable fatty acids, water, salts or electrolytes, such as prolaminesulfate, disodium hydrogen phosphate, potassium hydrogen phosphate,sodium chloride, zinc salts, colloidal silica, magnesium trisilicate,polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol,sodium carboxymethylcellulose, polyacrylates, waxes,polyethylene-polyoxypropylene-block polymers, polyethylene glycol andwool fat.

The compositions used in methods of treatment of the present invention,and pharmaceutical compositions of the invention, may be administeredorally, parenterally, by inhalation spray, topically, rectally, nasally,buccally, vaginally or via an implanted reservoir. The term “parenteral”as used herein includes subcutaneous, intravenous, intramuscular,intra-articular, intra-synovial, intrasternal, intrathecal,intrahepatic, intralesional and intracranial injection or infusiontechniques. Preferably, the compositions are administered orally,intraperitoneally or intravenously.

Sterile injectable forms of the compositions used in methods oftreatment of the present invention may be aqueous or oleaginoussuspension. These suspensions may be formulated according to techniquesknown in the art using suitable dispersing or wetting agents andsuspending agents. The sterile injectable preparation may also be asterile injectable solution or suspension in a non-toxicparenterally-acceptable diluent or solvent, for example as a solution in1, 3-butanediol. Among the acceptable vehicles and solvents that may beemployed are water, Ringer's solution and isotonic sodium chloridesolution. In addition, sterile, fixed oils are conventionally employedas a solvent or suspending medium. For this purpose, any bland fixed oilmay be employed including synthetic mono- or di-glycerides. Fatty acids,such as oleic acid and its glyceride derivatives are useful in thepreparation of injectables, as are natural pharmaceutically-acceptableoils, such as olive oil or castor oil, especially in theirpolyoxyethylated versions. These oil solutions or suspensions may alsocontain a long-chain alcohol diluent or dispersant, such as Ph. Helv orsimilar alcohol.

The pharmaceutical compositions of this invention may be orallyadministered in any orally acceptable dosage form including, but notlimited to, capsules, tablets, aqueous suspensions or solutions. In thecase of tablets for oral use, carriers which are commonly used includelactose and corn starch. Lubricating agents, such as magnesium stearate,are also typically added. For oral administration in a capsule form,useful diluents include lactose and dried corn starch. When aqueoussuspensions are required for oral use, the active ingredient is combinedwith emulsifying and suspending agents. If desired, certain sweetening,flavoring or coloring agents may also be added.

Alternatively, the pharmaceutical compositions of this invention may beadministered in the form of suppositories for rectal administration.These can be prepared by mixing the agent with a suitable non-irritatingexcipient which is solid at room temperature but liquid at rectaltemperature and therefore will melt in the rectum to release the drug.Such materials include cocoa butter, beeswax and polyethylene glycols.

The pharmaceutical compositions of this invention may also beadministered topically, especially to treat skin cancers, psoriasis orother diseases which occur in or on the skin. Suitable topicalformulations are readily prepared for each of these areas or organs.Topical application for the lower intestinal tract can be effected in arectal suppository formulation (see above) or in a suitable enemaformulation. Topically-acceptable transdermal patches may also be used.

For topical applications, the pharmaceutical compositions may beformulated in a suitable ointment containing the active componentsuspended or dissolved in one or more carriers. Carriers for topicaladministration of the compounds of this invention include, but are notlimited to, mineral oil, liquid petrolatum, white petrolatum, propyleneglycol, polyoxyethylene, polyoxypropylene compound, emulsifying wax andwater.

Alternatively, the pharmaceutical compositions can be formulated in asuitable lotion or cream containing the active components suspended ordissolved in one or more pharmaceutically acceptable carriers. Suitablecarriers include, but are not limited to, mineral oil, sorbitanmonostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol,2-octyldodecanol, benzyl alcohol and water.

For ophthalmic use, the pharmaceutical compositions may be formulated asmicronized suspensions in isotonic, pH adjusted sterile saline, or,preferably, as solutions in isotonic, pH adjusted sterile saline, eitherwith our without a preservative such as benzylalkonium chloride.Alternatively, for ophthalmic uses, the pharmaceutical compositions maybe formulated in an ointment such as petrolatum.

The pharmaceutical compositions of this invention may also beadministered by nasal aerosol or inhalation. Such compositions areprepared according to techniques well-known in the art of pharmaceuticalformulation and may be prepared as solutions in saline, employing benzylalcohol or other suitable preservatives, absorption promoters to enhancebioavailability, fluorocarbons, and/or other conventional solubilizingor dispersing agents.

The amount of compound in a pharmaceutical composition of the instantinvention that may be combined with the carrier materials to produce asingle dosage form will vary depending upon the host and diseasetreated, the particular mode of administration. Preferably, thecompositions should be formulated to contain between about 0.05milligram to about 750 milligrams or more (up to several grams), morepreferably about 1 milligram to about 600 milligrams, and even morepreferably about 10 milligrams to about 500 milligrams of activeingredient, alone or in combination with at least one additionalnon-antibody attracting compound which may be used to treat diabetes ora diabetes related disorder or to impact the effects of aging asotherwise set forth herein.

It should also be understood that a specific dosage and treatmentregimen for any particular patient will depend upon a variety offactors, including the activity of the specific compound employed, theage, body weight, general health, sex, diet, time of administration,rate of excretion, drug combination, and the judgment of the treatingphysician and the severity of the particular disease or condition beingtreated.

A patient or subject (e.g. a male human) suffering from or at risk ofdeveloping diabetes and/or a diabetes related disorder or who wishes toeffect aging processes can be treated by administering to the patient(subject) an effective amount of glucosepane and related aspects andembodiments according to the present invention includingpharmaceutically acceptable salts, solvates or polymorphs, thereofoptionally in a pharmaceutically acceptable carrier or diluent, eitheralone, or in combination with other known pharmaceutical agents,preferably agents which can assist in treating diabetes or a diabetesrelated disorder to ameliorate the secondary effects and conditionsassociated with diabetes. This treatment can also be administered inconjunction with other conventional therapies.

These compounds can be administered by any appropriate route, forexample, orally, parenterally, intravenously, intradermally,subcutaneously, or topically, in liquid, cream, gel, or solid form, orby aerosol form.

The active compound is included in the pharmaceutically acceptablecarrier or diluent in an amount sufficient to deliver to a patient atherapeutically effective amount for the desired indication, withoutcausing serious toxic effects in the patient treated. A preferred doseof the active compound for all of the herein-mentioned conditions is inthe range from about 10 ng/kg to 300 mg/kg, preferably 0.1 to 100 mg/kgper day, more generally 0.5 to about 25 mg per kilogram body weight ofthe recipient/patient per day. A typical topical dosage will range from0.01-3% wt/wt in a suitable carrier.

The compound is conveniently administered in any suitable unit dosageform, including but not limited to one containing less than 1 mg, 1 mgto 3000 mg, preferably 5 to 500 mg of active ingredient per unit dosageform. An oral dosage of about 25-250 mg is often convenient.

The active ingredient as described herein is preferably administered toachieve peak plasma concentrations of the active compound of about0.00001-30 mM, preferably about 0.1-30 μM. This may be achieved, forexample, by the intravenous injection of a solution or formulation ofthe active ingredient, optionally in saline, or an aqueous medium oradministered as a bolus of the active ingredient. Oral administration isalso appropriate to generate effective plasma concentrations of activeagent, as are topically administered compositions.

The concentration of active compound in the drug composition will dependon absorption, distribution, inactivation, and excretion rates of thedrug as well as other factors known to those of skill in the art. It isto be noted that dosage values will also vary with the severity of thecondition to be alleviated. It is to be further understood that for anyparticular subject, specific dosage regimens should be adjusted overtime according to the individual need and the professional judgment ofthe person administering or supervising the administration of thecompositions, and that the concentration ranges set forth herein areexemplary only and are not intended to limit the scope or practice ofthe claimed composition. The active ingredient may be administered atonce, or may be divided into a number of smaller doses to beadministered at varying intervals of time.

Oral compositions will generally include an inert diluent or an ediblecarrier. They may be enclosed in gelatin capsules or compressed intotablets. For the purpose of oral therapeutic administration, the activecompound or its prodrug derivative can be incorporated with excipientsand used in the form of tablets, troches, or capsules. Pharmaceuticallycompatible binding agents, and/or adjuvant materials can be included aspart of the composition.

The tablets, pills, capsules, troches and the like can contain any ofthe following ingredients, or compounds of a similar nature: a bindersuch as microcrystalline cellulose, gum tragacanth or gelatin; anexcipient such as starch or lactose, a dispersing agent such as alginicacid, Primogel, or corn starch; a lubricant such as magnesium stearateor Sterotes; a glidant such as colloidal silicon dioxide; a sweeteningagent such as sucrose or saccharin; or a flavoring agent such aspeppermint, methyl salicylate, or orange flavoring. When the dosage unitform is a capsule, it can contain, in addition to material of the abovetype, a liquid carrier such as a fatty oil. In addition, dosage unitforms can contain various other materials which modify the physical formof the dosage unit, for example, coatings of sugar, shellac, or entericagents.

The active compound or pharmaceutically acceptable salt thereof can beadministered as a component of an elixir, suspension, syrup, wafer,chewing gum or the like. A syrup may contain, in addition to the activecompounds, sucrose as a sweetening agent and certain preservatives, dyesand colorings and flavors.

The active compound or pharmaceutically acceptable salts thereof canalso be mixed with other active materials that do not impair the desiredaction, or with materials that supplement the desired action, such asanti-coagulant or coagulation inhibitory agents, anti-platelet orplatelet inhibitory agents, thrombin inhibitors, thrombolytic orfibrinolytic agents, anti-arrhythmic agents, anti-hypertensive agents,calcium channel blockers (L-type and T-type), cardiac glycosides,diuretics, mineralocorticoid receptor antagonists, phosphodiesteraseinhibitors, cholesterol/lipid lowering agents and lipid profiletherapies, traditional anti-diabetic agents, anti-depressants,anti-inflammatory agents (steroidal and non-steroidal),anti-osteoporosis agents, hormone replacement therapies, oralcontraceptives, anti-obesity agents, anti-anxiety agents,anti-proliferative agents, anti-tumor agents, anti-ulcer andgastroesophageal reflux disease agents, growth hormone and/or growthhormone secretagogues, thyroid mimetics (including thyroid receptorantagonist), antiinflammator agents, anticancer agents andanti-infective agents such as antibiotics, antifungals or antiviralcompounds, among others.

Solutions or suspensions used for parenteral, intradermal, subcutaneous,or topical application can include the following components: a sterilediluent such as water for injection, saline solution, fixed oils,polyethylene glycols, glycerine, propylene glycol or other syntheticsolvents; antibacterial agents such as benzyl alcohol or methylparabens; antioxidants such as ascorbic acid or sodium bisulfite;chelating agents such as ethylenediaminetetraacetic acid; buffers suchas acetates, citrates or phosphates and agents for the adjustment oftonicity such as sodium chloride or dextrose. The parental preparationcan be enclosed in ampoules, disposable syringes or multiple dose vialsmade of glass or plastic.

If administered intravenously, preferred carriers are physiologicalsaline or phosphate buffered saline (PBS).

In one embodiment, the active compounds are prepared with carriers thatwill protect the compound against rapid elimination from the body, suchas a controlled release formulation, including implants andmicroencapsulated delivery systems. Biodegradable, biocompatiblepolymers can be used, such as ethylene vinyl acetate, polyanhydrides,polyglycolic acid, collagen, polyorthoesters, and polylactic acid.Methods for preparation of such formulations will be apparent to thoseskilled in the art.

Liposomal suspensions may also be pharmaceutically acceptable carriers.These may be prepared according to methods known to those skilled in theart, for example, as described in U.S. Pat. No. 4,522,811 (which isincorporated herein by reference in its entirety). For example, liposomeformulations may be prepared by dissolving appropriate lipid(s) (such asstearoyl phosphatidyl ethanolamine, stearoyl phosphatidyl choline,arachadoyl phosphatidyl choline, and cholesterol) in an inorganicsolvent that is then evaporated, leaving behind a thin film of driedlipid on the surface of the container. An aqueous solution of the activecompound is then introduced into the container. The container is thenswirled by hand to free lipid material from the sides of the containerand to disperse lipid aggregates, thereby forming the liposomalsuspension.

Exemplary Processes and Compounds of the Invention—Establishing a Routeto an Efficient Chemical Synthesis of Glucosepane

In analyzing the glucosepane core, the inventors were first intrigued byits reported tendency to adopt an iso-imidazole topology, rather thanthat of the corresponding aromatic imidazole. The inventors thereforeset out to investigate this tautomeric preference through Gaussiancalculations performed on model compounds (Table 1).

TABLE 1 Results from density functional theory (DFT) calculationscomparing energies of various imidazole tautomerization states.

Entry X Y ΔG (kcal/mol)^(a) 1 H H 26.7 2 H NHMe 10.5 3 NMe₂ H −1.5 4NMe₂ NHMe −14.8 ^(a)All calculations were performed using the Gaussian09 program suite using the CBS-QB3 method. All calculations implementeda continuum model to account for the effects of water solvent.

While unsubstituted imidazole (Entry 1) greatly prefers the aromaticarrangement (ΔG=26.7 kcal/mol), addition of methylamino substituents tothe 2 and 4-position decreases this preference substantially (Entries2-3; ΔG=10.5 and −1.5 kcal/mol, respectively). Interestingly,2,4-diamino-substituted derivative (entry 4), which contains the samesubstitution pattern as glucosepane, exhibits a strong preference forthe non-aromatic tautomer. (ΔG=−14.8 kcal/mol). This trend may be partlyexplained by the decreasing aromatic stabilization of imidazole uponaddition of electron-donating substituents to the 2- and 4-positions, asindicated in prior work and additional calculations provided herein inthe Supporting Information.^((19),(20)) Furthermore, the inability ofelectron-donating substituents at the 2- and 4-positions to delocalizeinto the imidazole ring may drive a decrease in stabilization energy, aswell as a tendency to tautomerize into the iso-imidazole, which permitssuch delocalization. This model is further supported by geometryminimization experiments, which demonstrate for the 2,4-diaminoimidazole system that the 4-substituent is rotated such that N-lonepairs are partially non-overlapping with the heterocycle's pi-system.Taken together, the inventors hypothesized that in the setting of 4- and2,4-diamino imidazoles, the decrease in aromaticity does not afford asufficiently high degree of energetic stabilization; in theiso-imidazole tautomer, on the other hand, electron-donating amines havethe opportunity to participate extensively in resonance stabilization.

With this information in mind, the inventors constructed aretrosynthesis (FIG. 2). They reasoned that given the strongthermodynamics driving the core heterocycle's tautomerization state,formation of the C—N bonds between the arginine guanidine and thelysine-derived azepane (6+Arg→5) would be accompanied by spontaneousisomerization to the correct structure. They therefore first chose todisconnect at the two C—N bonds endocyclic to the imidazole motif. Thisdisconnection is identical to that suggested by Lederer and colleaguesfor the final step in the biosynthesis of glucosepane,⁽¹⁷⁾ whereinarginine is proposed to condense directly with an alpha-keto iminiumintermediate (6), formed from an adduct derived from lysine and glucose.They then reasoned that 6 could be generated through N-oxidation andregioselective elimination of azepane 7. In turn, azepane 7 could bedeconstructed via an Amadori rearrangement sequence to a suitablyprotected lysine derivative and known epoxide 8.⁽²¹⁾ In this sense, 8would serve as the source of the chiral diol encountered in glucosepanediastereomer 5. As it is unknown which stereoisomer(s) of glucosepaneare most prevalent in vivo, 8 was chosen as it reflects thestereochemistry of glucose, which is the most common precursor in vivo.In future studies, simply inverting the C-6 and C-7 stereocenters of thestarting epoxide 8 would then permit access to other reporteddiastereomers.

In the present invention (scheme 1, FIG. 3), the synthesis began withepoxide 8, prepared from diacetone-D-glucose as previouslydescribed.⁽²²⁾ A scheme showing the synthesis of epoxide 8 is presentedat the beginning of the experimental section. Nucleophilic addition ofDod-protected lysine derivative 9 to the less substituted side of theepoxide in 8, followed by acidic deprotection of the resulting tertiaryamine, provides amino alcohol 10 in 80% yield over two steps. Acetonideremoval in the presence of aqueous acetic acid then affords azepaneacetal 13. The conversion of 10 to 13 proceeds by way of intramolecularattack of the lysine amino group onto the anomeric carbon of thecarbohydrate with accompanying acetonide loss to give intermediateazepane hemiaminal 11.⁽²³⁾ This material spontaneously undergoes Amadorirearrangement^((24, 25)) to give an intermediate ketone 12, which isthen trapped intramolecularly by the C-6 hydroxyl to yield bridgedbicyclic acetal 13 in 60% yield. Reinstallation of the acetonide groupproceeds with reformation of the ketone functionality to afford thedesired protected ketone (14).

Access to 14 set the stage for oxidation-trapping attempts, outlined inthe retrosynthesis. Although the inventors were able to achieve thedesired α-keto iminium intermediate (16) by way of oxidation withSelectfluor®, this material rapidly undergoes ring contraction to givealdehyde 17 (Scheme 2A). All attempts to condense 17 with guanidinederivatives, including various protected forms of arginine, were metonly by complete decomposition of 17, and recovery of the guanidinenucleophile. Furthermore, attempts to perform oxidation and guanidinetrapping in “one pot” were also unsuccessful, providing similar resultsto the two-step process.

In light of these observations, the inventors decided to re-engineer theretrosynthesis. While the inventors were encouraged by their ability toaccess α-keto iminium 16, the refractoriness of this species tointermolecular trapping suggested that perhaps condensation with thearginine guanidine functionality succeeds in vivo because of proximityeffects. In other words, crosslinking is only likely to occur forproteins wherein an appropriately modified lysine residue is directlyadjacent to the attacking arginine, rendering the process functionallyintramolecular (even for proteins such as collagen whereinintermolecular crosslinking is accelerated due to the high localconcentration of reactive side-chains).^((3, 26)) In this light, itoccurred to the inventors that for the reaction to be successful, theymust tether the nucleophilic (guanidine) and electrophilic (iminium)components together at the time of oxidation.

In this light, they recognized that an intramolecular oxidation transferprocess—by way of a [3,3]-sigmatropic rearrangement from semicarbazonetautomer 19—would afford an intermediate (20) with the same coreoxidation state to α-keto iminium 16 (FIG. 4, Scheme 2B). Furthermore,20 also contains a tethered guanidine function perfectly poised forsubsequent intramolecular cyclization and tautomerization to afford theglucosepane core. Furthermore, we reasoned that simple condensation oflysine-derived ketone 14 with semicarbazide derivatives (Scheme 2C)would permit rapid access to semicarbazone 19, capable of tautomerizingto the desired [3,3] rearrangement substrate (20). In this sense, 20would function as a masked version of α-keto iminium 16, possessing thecorrect oxidation state and functional group disposition to afford thedesired iso-imidazole 18. Although such an imidazole-formation sequencehas never before been reported to the knowledge of the inventors, theywere encouraged by previous reports of analogous hetero-Claisenrearrangements.^((27, 30)) Taken together, the inventors envisioned thatthis sequence would accomplish the goal of directly coupling oxidationand condensation steps, solving the problems associated with ourintermolecular trapping sequence.

In the event (FIG. 5, Scheme 3A.), condensation of thiomethylsemicarbazide 22 with ketone 14 proceeded smoothly to affordsemicarbazone 24 (as a mixture of E/Z isomers) in 78% yield. Afterseveral failed attempts, the inventors were encouraged to discover thatupon treating semicarbazone 24 with excess chlorotrimethylsilane (TMSCl)in dry, refluxing chloroform induced the formation of iso-thioimidazole26. The inventors believe that this material forms by way of the pathwaypredicted in Scheme 2B, by way of tautomerization, [3,3]-sigmatropicrearrangement and cyclodeamination, and is accompanied by acetonideremoval, likely resulting from HCl generated by aqueous quenching ofexcess TMSCl. Notably, 26 is isolated as an epimeric mixture at C-8a, asconfirmed by NMR analysis.⁽³¹⁾ Interestingly, attempts to purifyiso-thioimidazole 26 under open atmosphere led to only to the isolationof C-8a-oxidized product 27.⁽³²⁾ By displacing the thiomethyl group withan ornithine derivative followed by C-8a reduction using Na(OAc)₃BH, theinventors were able to access protected glucosepane 28 (see SupportingInformation for details).

Despite this result, the inventors sought a more concise route tointermediate 28. Replacement of 22 in this sequence with a fullyelaborated amino-arginine derivative (23) readily afforded 25 in goodyield (69%), and gratifyingly, this intermediate also underwent thedesired rearrangement, cyclization and acetonide removal sequence. Thissequence therefore furnished fully protected glucosepane derivative 28in 4:1 diasteromeric ratio in only a single synthetic step.

With backbone-protected glucosepane in hand, completion of the syntheticsequence proved straightforward (FIG. 5, Scheme 3B.). Globalhydrogenolytic deprotection of Cbz and benzyl ester protecting groupswas achieved using Pd/C under an atmosphere of hydrogen gas and eitherTFA or formic acid (FIG. 5, Scheme 3B.), enabling rapid access to 5 aseither the formate or TFA salt. Although two C-8a epimers are producedin a 4:1 ratio through this route, these can be separated by preparativeHPLC. Spectral data obtained from ¹H- and ¹³C-NMR experiments usingsynthetic 5 proved identical to that reported by Lederer and colleaguesfor material isolated from model reactions.¹⁶ This confirms that thesynthetic glucosepane exists as the iso-imidazole tautomer. Overall,this synthetic route proceeds in a total of 8 steps and 12% overallyield and is the first to provide access to two of the eight possibleglucosepane diastereomers in enantiomerically pure form.

To provide further experimental evidence for 2,4-diaminoimidazoles toadopt the iso-imidazole tautomer, the inventors utilized our newlydeveloped rearrangement reaction to synthesize two additional2,4-diaminoimidazoles (29 and 30, FIG. 5, Scheme 3C.).⁽³³⁾ These wereprepared in two steps from commercially available starting materials,and as expected, were found to adopt the iso-imidazole tautomerexclusively. These results support our computational data, and confirmthat imidazoles with electron-rich substituents at the 2- and4-positions prefer the iso-imidazole tautomer.

With synthetic glucosepane (5) in hand, we next investigated itsstructural features using multidimensional NMR techniques.Interestingly, 2D ¹H-NMR NOESY experiments of compound 5 revealed thepresence of “conformational exchange” peaks,⁽³⁴⁾ which we attribute toE/Z isomerization about the exocyclic C2-N bond in glucosepane. Indeed,although the original glucosepane isolation report did note thepossibility of E/Z isomerism in acyclic 2-amino imidazoles, the presenceof these exchange peaks in the case of glucosepane have previously beenincorrectly attributed to stereoisomerism at the C-8a stereocenter.⁽³⁵⁾Using an EXSY (EXchange SpectroscopY) NOESY sequence,⁽³⁴⁾ the inventorswere able to calculate an approximate rate for this conformationalexchange process on the order of 3 s⁻¹ in D₂O.²⁸

The inventors next took advantage of glucospane's intrinsic spectralproperties to measure the pK_(a) of the iso-imidazole core. Theseexperiments revealed compound 5 to possess only one basic site underaqueous conditions with a pK_(a) of approximately 12, which theinventors believe to reflect protonation at the iso-imidazole N1 atom,consistent with both NMR data and DFT calculations.¹⁶ Exposure of eitherepimer of 5 to D₂O leads to quantitative hydrogen/deuterium exchange atthe C-8a H-atom, which occurs rapidly (<60 min) under basic conditions(aqueous NaOD). Taken together, these studies suggest that glucosepanecontains both acidic and basic sites, the latter of which possesses apK_(a) quite close to that of native arginine (pK_(a)=12.5). Therelevance of these structural features to glucosepanes biologicalproperties is currently being investigated in our laboratories.

Herein the inventors report the first total synthesis of glucosepane,which is rapid, high-yielding, and stereoselective. This contributionwas enabled by our development of a simple, one-pot protocol forsynthesizing several heavily substituted imidazoles via a sigmatropicrearrangement-cyclization sequence. This novel chemical process providesan entry point into other complex iso-imidazole-containing PTMs (e.g.,pentosinane), as well as various polyguanidine-containing naturalproducts. Furthermore, given that imidazoles are quite difficult toprepare in general, our one-pot synthetic route may prove useful as ageneral strategy for imidazole synthesis. The have performed detailedcomputational studies to explore the surprising tendency of substitutedimidazoles to adopt non-aromatic tautomerization states. These studiessuggest that imidazoles possessing electron-donating groups at the 2-and 4-positions possess a strong energetic preference for theiso-imidazole tautomer driven by effects of electron delocalization.Overall, the inventors believe that this data has significantimplications for understanding the molecular physiology ofglucosepane-protein adducts. Given that these are found in all humanbeings, and believed to be directly involved in the pathophysiologyunderlying various disease states, these results have the potential toprovide new directions for developing improved treatments for patients.More broadly, the inventors believe that the brevity and modularity ofthe disclosed synthesis will render it compatible with the site-specificincorporation ofglucosepane into synthetic oligopeptides, preparation ofaffinity reagents to identify molecular targets of glucosepane, thedevelopment of immunogens for raising antibodies against glucosepane,and also, perhaps, the identification of novel therapeutic strategiesfor glucosepane “crosslink-breaking.” Unlike prior investigations, whichhave been constrained to study glucosepane-modified proteins as highlyheterogeneous mixtures, the strategies reported herein have thepotential to open up entirely new opportunities for studying AGEs at anunprecedented level of resolution.

Experimental Details General Information for Chemical Synthesis

Starting materials were used as received unless otherwise noted. Allmoisture sensitive reactions were performed in an inert, dry atmosphereof nitrogen in oven dried glassware. Reagent grade solvents were usedfor extractions and flash chromatography. Reaction progress was checkedby analytical thin-layer chromatography (TLC, Merck silica gel 60 F-254plates). The plates were then monitored with UV illumination followed byvisualization with appropriate staining reagents such as anisaldehyde,ninhydrin, or KMnO₄. Flash column chromatography was performed usingsilica gel (230-400 mesh) using Teledyne Isco CombiFlash Rf200 unlessotherwise specified. The solvent compositions reported for allchromatographic separations are on a volume/volume (v/v) basis. Infrared(IR) spectra were recorded on a Thermo Nicolet 6700 FT-IR Spectrometer.¹H-NMR spectra were recorded on Agilent DD2 400 MHz, 500 MHz, 600 MHzand 800 MHz spectrometer and are reported in parts per million (ppm) onthe δ scale relative to CDCl₃ (δ 7.26), Methanol-d4 (δ 3.31), ACN-d3 (δ1.94), D₂O (δ 4.79) as an internal standard. Data are reported asfollows: chemical shift, multiplicity (s=singlet, d=doublet, t=triplet,q=quartet, br=broad, m=multiplet), coupling constants (Hz), andintegration. ¹³C-NMR spectra were recorded on Agilent DD2 125 MHz, and150 MHz spectrometers and are reported in parts per million (ppm) on theδ scale relative to CDCl₃ (δ 77.00), Methanol-d4 (δ 49.00), ACN-d3 (δ1.32). LC-MS analyses were performed on a Waters UPLC/S instrumentequipped with a RP-C18 column (1.7 μm particle size, 2.1×50 mm), dualatmospheric pressure chemical ionization (API)/electrospray (ESI) massspectrometry detector, and photodiode array detector. Optical rotationswere measured at 20° C.; concentrations are in g/100 mL.

Epoxide 8 Synthesis

O-((3aR,5R,6S,6aR)-5-((R)-2,2-dimethyl-1,3-dioxolan-4-yl)-2,2-dimethyltetrahydrofuro[2,3-d][1,3]dioxol-6-yl)S-methylcarbonodithioate (SI-2)

Diacetone-D-glucose (20 g, 76.8 mmol) was dissolved in dry THF (320 ml)at room temperature and 95% NaH (2.77 g, 115.3 mmol) was added in twoportions within 15 minutes with vigorous stirring and gas evolutionobserved. The reaction mixture was then stirred at room temp for 30minutes and to the resulting cloudy yellow solution was added CS₂ (9.28mL, 153.7 mmol) dropwise via syringe. This mixture was stirred for 30minutes and then MeI (8.13 mL, 130.6 mmol) was added dropwise viasyringe. Stirring was continued for another 30 minutes, at which pointTLC (1:1 Hexanes/EtOAc-KMnO₄ stain) indicated complete conversion. Theresulting brown solution, containing a white precipitate, was evaporatedin vacuo to a thick brown syrup, dissolved in EtOAc (300 mL) and washedwith water (300 mL). The aqueous layer was then back extracted twicewith EtOAc (200 mL) and the combined organic layers dried over MgSO₄,filtered and concentrated in vacuo to a thick dark orange oil.Purification was performed in 2 batches using Teledyne Isco with anormal phase 120 g RediSep column as the stationary phase. Hexanes andEtOAc were used as the mobile phase. Column conditions: 100% hexanes for2 column volumes (CVs) followed by 0→35% EtOAc over 14 CVs. The titlecompound was isolated in 99% yield (26.8 g, 76.75 mmol) as a thickyellow oil with spectra matching previously reported values.¹

(3aR,5S,6aR)-5-((R)-2,2-dimethyl-1,3-dioxolan-4-yl)-2,2-dimethyltetrahydrofuro[2,3-d][1,3]dioxole(SI-3)

Xanthante SI-2 (14.85 g, 42.4 mmol) was dissolved in dry toluene (210mL) and then n-Bu₃SnH (14.8 mL, 55.1 mmol) and freshly recrystallizedAIBN (1.4 g, 8.48 mmol) were added to the solution at room temperature.The reaction vessel was then fitted with a condenser and placed in anoil bath preheated to 90° C. Upon heating, gas evolution was observed.The reaction was stirred at 90° C. for 2 hours, at which point TLC (2:1Hexanes/EtOAc-KMnO₄ stain) indicated complete consumption of startingmaterial. The reaction was then cooled and concentrated in vacuo to givea pale yellow oil. Purification was performed using Teledyne Isco with anormal phase 120 g RediSep column as the stationary phase. Hexanes andEtOAc were used as the mobile phase. Column conditions: 100% hexanes for1 column volume (CV) followed by 0→5% EtOAc over 1 CV, then hold at 5%EtOAc for 2 CVs, followed by 5%→25% over 10 CVs, and finally hold at 25%EtOAc for 3 CVs. The title compound was isolated in 76% yield (8 g, 32.7mmol) as a thick yellow oil with spectra matching previously reportedvalues.¹

(R)-1-((3aR,5S,6aR)-2,2-dimethyltetrahydrofuro[2,3-d][1.3]dioxol-5-yl)ethane-1,2-diol(SI-4)

Acetal SI-3 (2.06 g, 8.46 mmol) was dissolved in 90% aqueous AcOH (6 mLH₂O in 54 mL glacial AcOH) and the reaction vessel was then placed in anoil bath preheated to 60° C. The reaction was stirred at thistemperature for 45 min (longer reaction times lead to second acetonidedeprotection), at which point TLC indicated complete consumption ofstarting material. The reaction mixture was then cooled to roomtemperature and concentrated in vacuo. Residual AcOH and water removedby azeotropic evaporation with toluene, and the resulting crude materialwas placed under high vacuum overnight to give a white solid.Purification was performed using Teledyne Isco with a normal phase 24 gRediSep column as the stationary phase. CH₂Cl₂ and MeOH were used as themobile phase. Column conditions: 100% CH₂Cl₂ for 1 column volume (CV)followed by 0→15% MeOH over 20 CVs. The title compound was isolated in93% yield (1.60 g, 7.83 mmol) as a white solid with spectra matchingpreviously reported values.¹

(R)-2-((3aR,5S,6aR)-2,2-dimethyltetrahydrofuro[2,3-d][1,3]dioxol-5-yl)-2-hydroxyethyl4-methylbenzenesulfonate (SI-5)

Diol SI-4 (1.08 g, 5.29 mmol) was dissolved in anhydrous pyridine (26mL) and cooled to 0° C. before freshly recrystallized tosyl chloride(1.16 g, 6.08 mmol) was added in one portion. The reaction mixture wasthen stirred at 0° C. while monitoring by TLC (1:1 Hex/EtOAc-KMnO₄stain). Generally, after 8 hrs the reaction showed complete consumptionof starting diol and formation of desired monotosylate as the majorproducts as well as the bis tosylate (ratio˜5:1—determined by ¹H NMR ofthe crude reaction mixture). Upon completion, the solvent was evaporatedin vacuo and the resulting crude residue purified by chromatography.Purification was performed using Teledyne Isco with a normal phase 40 gRediSep column as the stationary phase. Hexanes and EtOAc were used asthe mobile phase. Column conditions: 95% hexanes for 1 column volume(CV) followed by 5→50% EtOAc over 15 CVs, then 50% EtOAc for 2 CVsfollowed by 50→75% EtOAc over 2 CVs. The pure tosylate was isolated in84% yield (1.60 g, 4.46 mmol) as a colorless viscous oil with spectramatching previously reported values.²

(3aR,5S,6aR)-2,2-dimethyl-5-((R)-oxiran-2-yl)tetrahydrofuro[2,3-d][1,3]dioxole(8)

Tosylate SI-5 (1.55 g, 4.32 mmol) was dissolved in anhydrous MeOH (30mL) and the reaction mixture cooled to 0° C. NaOMe (1.16 g, 21.6 mmol)was then added in one portion. The reaction was stirred at 0° C. for 30min and then allowed to warm to room temperature, at which point TLC(2:1 Hex/EtOAc—KMnO₄ stain) showed complete conversion of startingmaterial to desired product. Approximately 80% of the solvent volume wasthen removed in vacuo and the resulting mixture was partitioned betweenequal volumes (30 mL) of water and Et₂O. The organic layer was collectedand the aqueous layer re-extracted with Et₂O (50 mL). The combined Et₂Olayers were dried over Mg₂SO₄, filtered, and concentrated in vacuo toafford a pale yellow oil (0.662 g, 3.58 mmol, 83% yield), which was usedwithout further purification. Spectra matched previously reportedvalues.³

Ketoazepane 14 Synthesis:

Benzyl N²-((benzyloxy)carbonyl)-N⁶-(bis(4-methoxyphenyl)methyl)-L-lysine(9)

Bis(4-methoxyphenyl)methanol (DodOH) (8.00 g, 32.7 mmol) was added to aflask containing Et₂O (200 mL), and the solution was cooled to 0° C. 12Maqueous HCl (16.3 mL, 196 mmol) was then added dropwise, which causedthe solution to turn slightly pink. The heterogeneous reaction mixturewas warmed to room temperature, stirred vigorously for 30 minutes, andthen poured into a separatory funnel. After separation of layers, theorganic layer was dried with a 1:1 mixture of MgSO₄ and NaHCO₃. Thesolid particles were filtered to give DodCl in Et₂O solution, which wasimmediately used in the next step without further purification.

Finely powdered Lysine-derived benzenesulfonte salt (13.3 g, 25.16 mmol)was added into a flask containing CH₂Cl₂ (500 mL). After cooling thesuspension to 0° C., Et₃N (17.5 mL, 125.7 mmol) was added. DodClsolution, freshly prepared as described above, was then added dropwiseover 30 minutes, which caused the solution to turn clear. The reactionmixture was warmed to room temperature, stirred for an additional 2hours, and then concentrated under vacuum to give crude material asyellow oil. Purification was performed in 2 batches using Teledyne Iscowith a normal phase 120 g RediSep column as the stationary phase. CH₂Cl₂and EtOAc were used as the mobile phase. Column conditions: 100% CH₂Cl₂for 1 column volume (CV) followed by 0→50% EtOAc over 15 CVs. Fullyprotected lysine derivative 9 was isolated in 88% yield (13.2 g, 22.14mmol) as thick oil. ¹H NMR (400 MHz, Chloroform-d) δ 7.37-7.29 (m, 10H),7.26 (d, J=8.6 Hz, 4H), 6.82 (d, J=8.5 Hz, 4H), 5.28 (d, J=8.3 Hz, 1H),5.16 (dd, J=19.0, 12.3 Hz, 2H), 5.09 (s, 2H), 4.68 (s, 1H), 4.41 (td,J=7.8, 5.1 Hz, 1H), 3.76 (s, 6H), 2.48 (t, J=6.9 Hz, 2H), 1.82 (dq,J=15.6, 5.3 Hz, 1H), 1.65 (ddt, J=18.4, 13.4, 5.9 Hz, 1H), 1.54-1.22 (m,4H). ¹³C NMR (101 MHz, cdcl₃) δ 172.25, 158.32, 155.76, 136.61, 136.11,135.19, 128.48, 128.40, 128.32, 128.14, 128.03, 127.98, 113.66, 66.94,66.85, 66.13, 55.09, 53.77, 47.65, 32.39, 29.61, 22.81.

HR-MS: (M+1)⁺=597.2967 (experimental); exact mass=597.2965 (theoretical)

[α]_(D)=−0.013° (c=5.1, CHCl₃)

BenzylN²-((benzyloxy)carbonyl)-N⁶-(bis(4-methoxyphenyl)methyl)-N⁶—((R)-2-((3aR,5S,6aR)-2,2-dimethyltetrahydrofuro[2,3-d][1,3]dioxol-5-yl)-2-hydroxyethyl)-L-lysine(SI-6)

Epoxide 8 (0.662 g, 3.56 mmol) and Dod lysine 9 (2.02 g, 3.39 mmol) weredissolved in iPrOH (17 mL), and the mixture was heated to reflux for 48hours, at which point TLC (1:1 Hex/EtOAc—CAM stain) indicated completeconversion. The solvent was removed in vacuo and the resulting cruderesidue purified by chromatography. Purification was performed usingTeledyne Isco with a normal phase 80 g RediSep column as the stationaryphase. Hexanes and EtOAc were used as the mobile phase. Columnconditions: 35% EtOAc for 1 column volume (V) followed by 35→55% EtOAcover 11 CVs. Pure SI-6 was isolated in 88% yield (2.45 g, 3.13 mmol) asa white foam.

¹H NMR (400 MHz, CDCl₃) δ 7.36-7.29 (m, 7H), 7.28-7.14 (m, 4H), 6.82(dd, J=10.8, 8.6 Hz, 3H), 5.77 (d, J=3.7 Hz, 1H), 5.26 (d, J=8.4 Hz,1H), 5.18-5.07 (m, 2H), 4.81 (s, 1H), 4.67 (t, J=4.2 Hz, 2H), 4.37 (td,J=8.0, 5.2 Hz, 1H), 3.97 (dt, J=10.0, 4.7 Hz, 1H), 3.78 (s, 7H), 3.76(s, 3H), 3.70 (dt, J=9.2, 4.8 Hz, 1H), 3.32 (s, 1H), 2.59 (dd, J=13.1,4.0 Hz, 1H), 2.54-2.33 (m, 3H), 1.91 (dd, J=13.5, 4.5 Hz, 1H), 1.75(ddd, J=13.4, 10.4, 4.8 Hz, 2H), 1.65-1.53 (m, 2H), 1.47 (s, 3H),1.46-1.35 (m, 1H), 1.29 (s, 3H), 1.14 (dtd, J=15.7, 10.3, 5.4 Hz, 2H).¹³C NMR (151 MHz, CDCl₃) δ 172.32, 158.68, 158.60, 155.92, 136.29,135.35, 133.90, 132.99, 129.87, 129.54, 128.68, 128.59, 128.54, 128.36,128.24, 128.22, 128.17, 113.85, 113.77, 111.08, 105.46, 80.46, 80.07,69.15, 68.28, 67.17, 67.05, 55.27, 55.26, 53.85, 53.47, 50.81, 33.71,32.57, 26.82, 26.45, 26.19, 22.97.

HR-MS: (M+1)⁺=783.3834 (experimental); exact mass=783.3857 (theoretical)

IR f (cm⁻¹): 1718, 1609, 1508, 1455, 1242, 1165, 1019, 822, 736, 697.

[α]_(D)=+0.028° (c=4.3. CHCl₃)

BenzylN²-((benzyloxy)carbonyl)-N⁶—((R)-2-((3aR,5S,6aR)-2,2-dimethyltetrahydrofuro[2,3-d][1,3]dioxol-5-yl)-2-hydroxyethyl)-L-lysinate2,2,2-trifluoroacetate (10)

Dod lysine derivative SI-6 (1.00 g, 1.28 mmol) was dissolved inanhydrous CH₂Cl₂ (12.35 mL) and cooled to 0° C. before TIPS (1.3 mL,6.39 mmol), anisole (0.208 mL, 1.92 mmol) and TFA (0.650 mL) weresequentially added to the cooled reaction and allowed to stir at 0° C.while monitoring by TLC (1:2 Hex/EtOAc, CAM stain—a 10 □| aliquot wasquenched with aqueous NaHCO₃ before spotting on the TLC plate.) Uponcompletion (approx. 6 h) the reaction was quenched at 0° C. with aqueousNaHCO₃ (20 mL) and the mixture transferred to a separatory funnel withthe aid of CH₂Cl₂ (20 mL). The organic layer was drawn off and theaqueous layer was then re-extracted with two portions of CH₂Cl₂ (20 mL).The combined organic extracts were dried over anhydrous Mg₂SO₄,filtered, and evaporated to dryness. The resulting crude residue waspurified by chromatography. Purification was performed using TeledyneIsco with a normal phase 12 g RediSep column as the stationary phase.CH₂Cl₂ and MeOH were used as the mobile phase. Column conditions: 100%CH₂Cl₂ for 3 column volumes (CVs) followed by 0→15% MeOH over 20 CVs,then 15% MeOH for 7 CVs. Pure secondary amine-TFA salt 10 was isolatedin 91% yield (0.781 g, 1.16 mmol) as a pale yellow viscous oil.

¹H NMR (600 MHz, CDCl₃) δ 7.38-7.26 (m, 10H), 5.75 (d, J=3.6 Hz, 1H),5.56 (d, J=8.2 Hz, 1H), 5.16 (q, J=12.2 Hz, 2H), 5.07 (t J=10.8 Hz, 2H),4.69 (t, J=4.1 Hz, 1H), 4.37 (tt, J=8.8, 3.5 Hz, 1H), 4.07 (ddd, J=10.7,6.4, 4.5 Hz, 1H), 3.96 (ddd, J=9.8, 6.3, 2.9 Hz, 1H), 3.10 (dd, J=12.6,2.9 Hz, 1H), 2.91 (dd, J=12.7, 9.9 Hz, 1H), 2.86 (t, J=7.8 Hz, 2H), 2.18(dd, J=13.5, 4.5 Hz, 1H), 1.84 (qd, J=10.8, 8.6, 3.2 Hz, 1H), 1.74-1.61(m, 4H), 1.47 (s, 3H), 1.35 (qd, J=10.9, 9.5, 4.9 Hz, 2H), 1.29 (s, 3H).¹³C NMR (151 MHz, CDCl₃) δ 172.12, 156.19, 136.31, 135.34, 128.77,128.67, 128.65, 128.51, 128.33, 128.26, 111.71, 105.73, 80.44, 79.23,68.69, 67.41, 67.20, 53.73, 50.95, 48.16, 35.42, 32.04, 26.84, 26.21,25.40, 22.35.

HR-MS: (M+1)+=557.2898 (experimental); exact mass=557.2863 (theoretical)

IR f (cm⁻¹) λ_(max): 1672, 1528, 1455, 1375, 1200, 1132, 1055, 1019,835, 799, 736, 721, 697.

[α]_(D)=−0.038° (c=5.4, CHCl₃)

Benzyl(S)-2-(((benzyloxy)carbonyl)amino)-6-((1R,5R,6S)-1,6-dihydroxy-8-oxa-3-azabicyclo[3.2.1]octan-3-yl)hexanoate(13)

Lysine derivative 10 (1.60 g, 2.39 mmol) was dissolved in 70% aqueousAcOH (24 mL) and the reaction mixture heated to 65° C. Upon completeconsumption of starting material (˜36 h), as determined by LCMS analysisof the crude reaction mixture, the solvent was removed in vacuo and theresulting brown residue dissolved in ACN (24 mL) and heated at 65° C.for 2 hrs. The reaction was then cooled and evaporated to dryness, andthe resulting residue was purified by chromatography. Purification wasperformed using Teledyne Isco with a normal phase 40 g RediSep column asthe stationary phase. Hexanes and EtOAc were used as the mobile phase.Column conditions: 70% EtOAc for 4 column volumes (CVs) followed by70→90% EtOAc over 16 CVs, 90% EtOAc for 6 CVs, 90→100% EtOAc over 1 CV,and finally 100% EtOAc over 7 CVs. Pure azepane acetal 13 was isolatedin 60% yield (0.715 g, 1.43 mmol) as a pale yellow viscous oil.

¹H NMR (500 MHz, MeOH-d4) δ 7.32 (dtt, J=19.0, 6.6, 3.1 Hz, 10H), 5.17(dd, J=31.6, 12.2 Hz, 2H), 5.09 (dd, J=20.5, 12.4 Hz, 2H), 4.23 (ddd,J=14.4, 8.3, 4.1 Hz, 2H), 4.01-3.96 (m, 2H), 2.64 (dd, J=11.0, 5.5 Hz,2H), 2.43 (dd, J=12.7, 7.4 Hz, 1H), 2.27 (t, J=6.9 Hz, 2H), 2.00 (dd,J=10.6, 7.4 Hz, 2H), 1.82 (dq, J=13.2, 7.1, 6.4 Hz, 1H), 1.68 (dq,J=14.2, 7.6, 6.9 Hz, 1H), 1.56 (ddt, J=12.4, 2.5, 1.0 Hz, 1H), 1.41 (dh,J=29.2, 7.8 Hz, 4H).

¹³C NMR (126 MHz, MeOH-d4) δ 174.00, 158.66, 138.16, 137.27, 129.56,129.45, 129.30, 129.24, 128.97, 128.77, 104.65, 83.27, 74.93, 67.79,67.64, 62.38, 58.04, 55.48, 46.59, 32.29, 26.78, 24.38.

HR-MS: (M+1)+=499.2444 (experimental); exact mass=499.2423 (theoretical)

IR f (cm⁻¹) λ_(max): 3324.5, 2947, 1717, 1527, 1454, 1393, 1341, 1260,1185, 1057, 954, 738, 698.

[α]_(D)=−0.83° (c=6.5, CHCl₃)

Benzyl(S)-2-(((benzyloxy)carbonyl)amino)-6-((3aR,8aS)-2,2-dimethyl-7-oxohexahydro-5H-[1,3]dioxolo[4,5-c]azepin-5-yl)hexanoate(14)

Acetal 13 (0.650 g, 1.30 mmol) was dissolved in anhydrous toluene (13mL), and PPTS (0.066 g, 0.26 mmol) and 2,2-dimethoxy propane (1.6 mL,13.0 mmol) were sequentially added. The resulting reaction mixture washeated to 95° C. and monitored by TLC (2:1 Hex/EtOAc) and LCMS. Uponcomplete consumption of starting material (˜18 h) the reaction mixturewas cooled to room temperature and partitioned between equal volumes (20mL) of CH₂Cl₂ and saturated aqueous NaHCO₃. The bottom organic layer wasdrawn off and the aqueous layer was then washed twice with CH₂Cl₂ (20mL). The combined organic layers were dried over anhydrous Na₂SO₄,filtered, and evaporated to dryness. The resulting dark brown residuewas purified by column chromatography using SiO₂ as stationary phase and3:2 Hexanes/EtOAc containing 2% NEt₃ as the mobile phase. The titlecompound was isolated in 85% yield (0.595 g, 1.11 mmol) as pale yellowviscous oil.

¹H NMR (600 MHz, CDCl₃) δ 7.39-7.30 (m, 10H), 5.30 (d, J=8.4 Hz, 1H),1.33-1.23 (m, 2H), 5.17 (dd, J=34.2, 12.0 Hz, 2H), 5.10 (s, 2H), 4.43(td, J=8.0, 5.2 Hz, 1H), 4.24 (ddd, J=6.5, 5.2, 3.8 Hz, 1H), 3.14 (dd,J=21.5, 17.0 Hz, 2H), 1.49-1.36 (m, 1H), 4.09 (dt, J=7.5, 5.0 Hz, 1H),3.00 (dd, J=11.4, 6.9 Hz, 1H), 2.97 (dd, J=11.3, 3.8 Hz, 1H), 2.70 (dd,J=14.9, 4.8 Hz, 1H), 2.60 (dd, J=14.7, 7.7 Hz, 2H), 2.47 (t, J=6.6 Hz,2H), 1.87 (ddt, J=14.7, 10.1, 5.0 Hz, 1H), 1.68 (dddd, J=13.7, 9.5, 7.2,5.8 Hz, 1H), 1.41 (s, 3H), 1.31 (s, 3H).

¹³C NMR (151 MHz, CDCl₃) 5208.97, 172.33, 155.93, 136.25, 135.33,128.72, 128.62, 128.39, 128.31, 128.22, 108.16, 76.06, 73.98, 68.28,67.21, 67.09, 57.42, 55.44, 53.90, 43.27, 32.53, 28.36, 27.29, 25.87,22.63.

HR-MS: (M+1)⁺=539.2757 (experimental); exact mass=539.2744 (theoretical)

IR f (cm⁻¹) □_(max): 2936, 1714, 1523, 1454, 1380, 1341, 1240, 1213,1046, 909, 861, 735, 697.

[α]_(D)=+0.0210 (c=4.9, CHCl₃)

Benzyl(S)-2-(((benzyloxy)carbonyl)amino)-6-((3aR,7aS)-6-formyl-2,2-dimethyl-3a,7a-dihydro-[1,3]dioxolo[4,5-c]pyridin-5(4H)-yl)hexanoate(17)

Ketone 14 (0.040 g, 0.0745 mmol) was dissolved in anhydrous ACN (0.750mL) under N₂, and the resulting solution was cooled to −40° C.Selectfluor (0.032 g, 0.0892 mmol) was added in one portion and theresulting reaction mixture was stirred at −40° C. for 2 hours. Thereaction was then quenched by addition of saturated aqueous NaHCO₃(0.300 mL) and resulting biphasic mixture was allowed to warm to roomtemp with vigorous stirring. Mixture was then partitioned between equalvolumes (5 mL) of saturated aqueous NaHCO₃ and CH₂Cl₂, bottom organiclayer drawn off and remaining aqueous layer extracted with CH₂Cl₂ (2×).The combined organic layers were dried over anhydrous Na₂SO₄, filtered,and evaporated to dryness. The resulting dark brown residue was purifiedby column chromatography using SiO₂ as stationary phase and 3:1Hexanes/EtOAc containing 1% NEt₃ as the mobile phase. The title compoundwas isolated in 80% yield (0.032 g, 0.596 mmol) as pale yellow viscousoil.

¹H NMR (600 MHz, CDCl₃) δ 9.11 (s, 1H), 7.34 (tt, J=10.8, 5.4 Hz, 10H),5.49 (d, J=4.1 Hz, 1H), 5.34 (d, J=8.4 Hz, 1H), 5.16 (q, J=12.3 Hz, 2H),5.10 (s, 2H), 4.63 (t, J=5.0 Hz, 1H), 4.40 (td, J=7.9, 5.2 Hz, 1H), 4.14(dt, J=8.9, 4.9 Hz, 1H), 3.33 (t, J=7.6 Hz, 2H), 3.12 (dd, J=12.9, 4.1Hz, 1H), 2.85 (dd, J=12.9, 8.0 Hz, 1H), 1.84 (ddt, J=15.8, 10.8, 5.3 Hz,1H), 1.73-1.61 (m, 1H), 1.50-1.19 (m, 4H), 1.44 (s, 3H), 1.39 (s, 3H).¹³C NMR (126 MHz, CDCl₃) δ 189.60, 172.19, 155.85, 145.98, 136.31,135.36, 128.63, 128.60, 128.49, 128.43, 128.34, 128.27, 128.16, 128.12,128.05, 127.91, 118.32, 108.72, 71.54, 69.63, 67.09, 66.98, 53.91,50.18, 50.12, 32.45, 28.52, 28.40, 26.00, 22.27.

HR-MS: (M+1)⁺=536.2530 (experimental); exact mass=536.2523 (theoretical)

IR f (cm⁻¹) ex: 2950, 1695, 1499, 1451, 1240, 1213, 1046, 909, 861, 801,693.

[α]_(D)=+0.017° (c=1.3, CHCl₃)

Iso-Imidazole Synthesis (Oxidation/Addition/Reduction Sequence):

Benzyl(S)-2-(((benzyloxy)carbonyl)amino)-6-((3aR,8aS,Z)-7-(2-(imino(methylthio)methyl)hydrazono)-2,2-dimethylhexahydro-5H-[1,3]dioxolo[4,5-c]azepin-5-yl)hexanoate(24)

Ketone 24 (0.560 g, 1.04 mmol) was dissolved in anhydrous MeOH (10 mL)and SMe-semicarbazide hydroiodide (0.267 g, 1.14 mmol) was added in oneportion at room temperature. The resulting reaction mixture was stirredand monitored by LCMS. Upon complete consumption of starting material(˜3 h), the mixture was evaporated to dryness to afford a orange-yellowfoam. This material was then portioned between equal volumes (10 mL) ofCH₂Cl₂ and 1 M NaOH and the resulting biphasic mixture stirredvigorously for 5 min. After separation of layers, the organic phase wasdrawn off and the aqueous layer re-extracted twice with CH₂Cl₂ (10 mL).The combined organic layers were dried over anhydrous Na₂SO₄, filtered,and concentrated to dryness. The resulting crude pale orange viscous oilwas purified by column chromatography using SiO₂ as stationary phase and2:3 Hexanes/EtOAc containing 3% NEt₃ as the mobile phase. The desiredsemicarbazone was isolated in 74% yield (0.482 g, 0.771 mmol) as a paleyellow viscous oil (1.5:1 mixture of E/Z isomers).

Minor isomer: ¹H NMR (600 MHz, Acetonitrile-d₃) δ 7.40-7.29 (m, 10H),6.01 (s, 1H), 5.64 (s, 1H), 5.13 (q, J=12.4 Hz, 2H), 5.06 (dd, J=14.4,12.5 Hz, 2H), 4.22-4.17 (m, 1H), 4.16-4.11 (m, 1H), 4.06 (q, J=5.9 Hz,1H), 3.66 (d, J=16.6 Hz, 1H), 3.40 (d, J=16.5 Hz, 1H), 2.82 (dd, J=12.5,8.4 Hz, 1H), 2.63 (dd, J=12.5, 3.8 Hz, 1H), 2.61 (d, J=1.5 Hz, 2H), 2.43(q, J=6.2 Hz, 2H), 2.34 (s, 3H), 1.80 (dt, J=15.5, 7.1 Hz, 1H),1.70-1.62 (m, 1H), 1.43-1.37 (m, 2H), 1.35 (s, 3H), 1.34-1.26 (m, 2H),1.26 (s, 3H).

Major isomer: ¹H NMR (600 MHz, Acetonitrile-d₃) δ 7.40-7.29 (m, 101H),6.01 (d, J=8.2 Hz, 1H), 5.66 (s, 1H), 5.13 (q, J=12.5 Hz, 2H), 5.06 (s,2H), 4.21-4.18 (m, 1H), 4.19-4.16 (m, 1H), 4.13 (q, J=6.2 Hz, 1H), 3.43(dd, J=11.4, 4.8 Hz, 1H), 3.24 (dd, J=29.8, 13.2 Hz, 2H), 2.78-2.75 (m,2H), 2.49-2.40 (m, 2H), 2.38 (s, 3H), 1.83-1.75 (m, 1H), 1.69-1.63 (m,1H), 1.49-1.39 (m, 2H), 1.36 (s, 3H), 1.31-1.25 (m, 2H), 1.24 (s, 3H).

Mixture of E/Z Isomers

¹³C NMR (151 MHz, Acetonitrile-d₃) δ 173.39, 161.90, 161.19, 159.85,159.68, 157.16, 138.10, 137.11, 129.50, 129.44, 129.17, 129.02, 128.91,128.90, 128.69, 108.57, 108.29, 76.95, 76.78, 74.32, 67.40, 67.10,63.50, 58.15, 58.11, 56.78, 56.63, 55.29, 55.26, 54.71, 38.14, 32.75,32.13, 32.10, 28.60, 28.12, 28.08, 27.99, 27.65, 25.90, 25.26, 25.19,23.94, 23.88, 12.73.

HR-MS: (M+1)⁺=626.3012 (experimental); exact mass=626.3016 (theoretical)

IR f (cm⁻¹): 2931, 1716, 1592, 1533, 1454, 1380, 1244, 1211, 1154, 1040,908, 734, 697.

[α]_(D)=+0.046° (c=1.2, CHCl₃)

Benzyl(2S)-2-(((benzyloxy)carbonyl)amino)-6-((5aR,8aS)-7,7-dimethyl-2-(methyltho)-5a,8a,9,9a-tetrahydro-[1,3]dioxolo[4,5-e]imidazo[4,5-b]azepin-4(5H)-yl)hexanoate(26)

Semicarbazone 24 (0.050 g, 0.0799 mmol) was dissolved in anhydrous anddegassed (freeze/pump/thaw) CHCl₃ (0.800 mL) before freshly distilledand degassed TMSCl (0.029 mL, 0.239 mmol) was added to the solution atroom temperature. The reaction vessel was sealed under argon and placedin an oil bath preheated to 95° C. The reaction was stirred at thistemperature for 24 h, at which point LCMS analysis of the resulting darkreddish-brown mixture indicated complete consumption of startingmaterial and formation of desired thioisoimidazole ([M+H]⁺ 609). Thereaction mixture was then treated with water (0.020 mL) and stirred atroom temperature for 1 hour. LCMS analysis indicated full deprotectionof the acetonide group ([M+H]⁺ 596). The reaction mixture was cooled inan ice bath and diluted with 1 M aqueous HCl (1.00 mL) and CH₂Cl₂ (0.300mL). The resulting biphasic mixture was stirred vigorously for 5 min.then layers were allowed to separate, the bottom organic layer drownoff, and the aqueous layer re-extracted twice with CH₂Cl₂ (1.00 mL). Thecombined organic layers were dried over anhydrous Na₂SO₄, filtered, andconcentrated to dryness. The title compound (0.052 g, crude mass) wasisolated as a HCl salt and carried on without further purification.

For characterization purposes the title compound was purified bypreparatory HPLC with a SunFire Prep C18 OBD 5 μm 10×150 mmreversed-phase column as the stationary phase. H₂O and MeCN bothbuffered with 0.1% trifluoroacetic acid were used as the mobile phase.HPLC conditions: UV collection 254 nm, flow rate 5 mL/min, 25%→60% MeCNlinear gradient over 32 minutes. The HPLC fractions were combined andlyophilized and title compound was isolated as a pale red fluffy solid.

Prep HPLC Retention Time: 12.72 min (minor isomer) & 13.18 min (majorisomer).

Due to rapid epimerization of the two isolated isomers, the titlecompound was characterized as a mixture of epimers.

¹H NMR (600 MHz, Acetonitrile-d₃) δ 7.37 (dq, J=11.8, 8.0 Hz, 20H), 6.03(d, J=8.4 Hz, 1H), 6.00 (d, J=8.3 Hz, 1H), 5.30 (dd, J=12.1, 2.4 Hz,1H), 5.18-5.12 (m, 4H), 5.11-5.05 (m, 4H), 4.86 (dd, J=12.3, 3.2 Hz,1H), 4.22 (qd, J=9.0, 4.9 Hz, 3H), 4.12-4.04 (m, 2H), 3.89 (dt, J=14.0,7.3 Hz, 1H), 3.82 (d, J=11.2 Hz, 1H), 3.77-3.63 (m, 4H), 3.56-3.41 (m,2H), 3.17 (d, J=14.5 Hz, 1H), 2.63 (s, 6H), 2.39 (dt, J=14.2, 3.9 Hz,1H), 2.20 (td, J=12.8, 3.9 Hz, 1H), 1.90-1.60 (m, 10H), 1.37 (pd, J=7.6,2.1 Hz, 4H). ¹³C NMR (151 MHz, Acetonitrile-d₃) 185.48, 185.45, 184.65,183.32, 173.24, 173.18, 157.23, 138.08, 137.10, 129.54, 129.48, 129.22,129.03, 128.96, 128.70, 72.18, 70.82, 70.29, 69.24, 67.50, 67.48, 67.18,67.16, 63.45, 61.80, 55.03, 54.99, 54.75, 53.55, 53.47, 51.75, 34.55,31.76, 31.72, 30.22, 26.96, 26.77, 23.16, 23.12, 14.70, 14.67.

HR-MS: (M+1)⁺=569.2440 (experimental); exact mass=569.2428 (theoretical)

IR f (cm⁻¹): 3209, 2938, 1677, 1642, 1521, 1431, 1365, 1202, 1136, 800,722, 699.

(6R,7S)-4-((S)-6-(benzyloxy)-5-(((benzyloxy)carbonyl)amino)-6-oxohexyl)-6,7,8a-trihydroxy-2-(methylthio)-4,5,6,7,8,8a-hexahydroimidazo[4,5-b]azepin-1-iumtrifluoroacetate (27)

Crude isothioimdazole 26 was dissolved in a 4:1 mixture of acetonitrile(0.640 mL) and DI water (0.160 mL) before NEt₃ (0.011 mL, 0.0799 mmol)was added. The resulting dark brown reaction mixture was stirred at roomtemperature under ambient air. LCMS analysis of the dark reddish-brownmixture indicated complete consumption of starting material (˜10 h) andformation of desired oxothioisoimidazole. This reaction mixture wascarried on to the subsequent addition step.

For characterization purposes the title compound was purified bypreparatory HPLC with a SunFire Prep C18 OBD 10 □m 19×150 mm reversedphase column as the stationary phase. H₂O and MeCN both buffered with0.1% trifluoroacetic acid were used as the mobile phase. HPLCconditions: UV collection 254 nm, flow rate 20 mL/min, 34%→45% MeCNlinear gradient over 30 minutes. The HPLC fractions were combined andlyophilized. The title compound isolated as a white solid.

Prep HPLC Retention Time: 6.92 min

¹H NMR (500 MHz, Acetonitrile-d₃) 7.35 (dd, J=9.7, 6.9 Hz, 10H), 6.03(d, J=8.2 Hz, 1H), 5.12 (dd, J=16.1, 12.6 Hz, 2H), 5.06 (dd, J=16.3,12.9 Hz, 2H), 4.23 (d, J=14.8 Hz, 1H), 4.19 (dd, J=8.5, 5.1 Hz, 1H),4.08-4.04 (m, 1H), 4.01 (dd, J=11.4, 3.7 Hz, 1H), 3.92 (dt, J=14.1, 7.4Hz, 1H), 3.52 (dd, J=14.8, 6.6 Hz, 1H), 3.41 (dt, J=13.5, 6.6 Hz, 1H),2.58 (s, 3H), 2.32 (dd, J=13.8, 4.1 Hz, 1H), 2.08-1.99 (m, 1H), 1.83(ddd, J=13.3, 8.2, 3.8 Hz, 1H), 1.67 (dtt, J=21.2, 14.0, 6.7 Hz, 3H),1.35 (p, J=7.6 Hz, 2H). ¹³C NMR (126 MHz, cd₃cn) δ 185.87, 180.20,173.22, 157.24, 138.09, 137.11, 129.54, 129.48, 129.21, 129.04, 128.95,128.70, 92.87, 70.52, 70.50, 67.49, 67.18, 55.65, 55.06, 52.34, 35.06,31.74, 26.72, 23.14, 14.55.

HR-MS: (M+1)⁺=585.2363 (experimental); exact mass=585.2377 (theoretical)

[α]_(D)=−0.018° (c=2.3, ACN-d4)

(6R,7S)-4-((S)-6-(benzyloxy)-5-(((benzyloxy)carbonyl)amino)-6-oxohexyl)-2-(((S)-4-(((benzyloxy)carbonyl)amino)-4-carboxybutyl)amino)-6,7,8a-trihydroxy-4,5,6,7,8,8a-hexahydroimidazo[4,5-b]azepin-1-ium(SI-8)

To the reaction mixture obtained in the previous step, Z-Orn-OH—HCl(0.029 g, 0.0959 mmol) was added to the reaction flask, followed by NEt₃(0.011 mL, 0.0799 mmol). The resulting reaction mixture was heated to50° C. and stirred until LCMS analysis of the reaction mixture indicatedcomplete consumption of starting material and formation of oxidizedadduct SI-8 (M+H−803.4). The reaction mixture was then cooled in an icebath and diluted with 1M aqueous HCl (1.0 mL) and CH₂Cl₂ (0.300 mL). Theresulting biphasic mixture was stirred vigorously for 5 min, then layerswere allowed to separate, the bottom organic layer drown off, and theaqueous re-extracted twice with CH₂Cl₂ (1.0 mL). The combined organiclayers were dried over anhydrous Na₂SO₄, filtered, and concentrated todryness. The title compound (0.080 g, crude mass) was isolated as an HClsalt in an undetermined mixture of epimers and carried on to the nextstep without further purification.

For characterization purposes the title compound was purified bypreparative HPLC. Preparatory HPLC was performed with a Macherey-NagelNucleosil Prep C18, 7 □m, 21×250 mm reversed-phase column as thestationary phase. H₂O and MeCN both buffered with 0.1% trifluoroaceticacid were used as the mobile phase. HPLC conditions: UV collection 254nm, flow rate 20 mL/min, 25%→45% MeCN linear gradient over 35 minutesand then isocratic elution with 45% MeCN over 5 min. The HPLC fractionswere combined and lyophilized.

Prep HPLC Retention Time: 39.57 min

¹H NMR (600 MHz, 15% D₂O, Acetonitrile-d₃) δ 7.39-7.26 (m, 15H),5.13-4.98 (m, 6H), 4.16 (dtd, J=26.4, 9.2, 5.3 Hz, 2H), 4.08-4.01 (m,1H), 3.99 (dt, J=6.8, 3.3 Hz, 1H), 3.93-3.73 (m, 3H), 3.45-3.36 (m, 2H),3.29-3.14 (m, 2H), 2.22 (ddd, J=18.9, 13.8, 4.4 Hz, 1H), 1.99-1.95 (m,1H), 1.88-1.76 (m, 2H), 1.71-1.54 (m, 6H), 1.32 (dtd, J=14.6, 7.9, 6.9,4.3 Hz, 2H). ¹³C NMR (151 MHz, 15% D₂O, Acetonitrile-d₃) δ 179.82,179.16, 178.82, 175.20, 174.98, 173.76, 173.73, 166.53, 165.97, 165.66,161.73, 161.61, 157.91, 157.82, 157.73, 157.71, 137.81, 137.78, 136.88,129.59, 129.57, 129.52, 129.50, 129.30, 129.27, 129.03, 129.00, 128.73,128.71, 128.69, 92.71, 92.44, 90.27, 89.49, 72.12, 71.80, 70.59, 70.34,70.31, 70.13, 70.11, 69.79, 67.73, 67.69, 67.48, 67.44, 67.38, 55.68,55.07, 54.99, 54.81, 54.73, 54.37, 54.29, 51.89, 51.83, 51.72, 42.77,42.55, 42.33, 40.10, 39.95, 35.70, 33.79, 31.56, 29.49, 29.33, 29.03,26.66, 26.52, 26.38, 25.62, 25.46, 23.17, 23.13.

HR-MS: (M+1)⁺=803.3595 (experimental); exact mass=803.3610 (theoretical)

IR f (cm⁻¹): 3294, 1681, 1610, 1533, 1426, 1345, 1203, 1139, 1055, 840,740, 723, 698.

[α]_(D)=−0.055° (c=2.7, ACN-d4)

2,2,2-trifluoro-1λ³-ethan-1-one,(6R,7S,Z)-4-((S)-6-(benzyloxy)-5-(((benzyloxy)carbonyl)amino)-6-oxohexyl)-2-(((S)-4-(((benzyloxy)carbonyl)amino)-4-carboxybutyl)-λ⁴-azanylidene)-6,7-dihydroxy-1,2,4,5,6,7,8,8a-octahydroimidazo[4,5-b]azepin-1-iumsalt (28)

Crude SI-8 was dissolved in anhydrous CHCl₃ (0.800 mL) and Na(AcO)₃BH(0.084 g, 0.400 mmol) was added in one portion. The resulting reactionmixture was stirred at room temperature for 15 hours. LCMS analysis ofthe dark reddish-brown mixture indicated complete consumption ofstarting material and formation of tittle compound ([M+H]⁺ 877). Thereaction was quenched by drop wise addition of 10% aqueous TFA (1 mL).The resulting biphasic mixture was stirred vigorously for 5 min, thenlayers were allowed to separate, the bottom layer drawn off, and theaqueous layer re-extracted twice with CH₂Cl₂ (1.0 mL). The combinedorganic layers were dried over anhydrous Na₂SO₄, filtered, andconcentrated to a dark brown residue containing a 4:1 mixture of epimers(0.078 g, crude mass) which was separated by preparative scale HPLC.Preparatory HPLC was performed with a Macherey-Nagel Nucleosil Prep C18,7 □m, 21×250 mm reversed-phase column as the stationary phase. H₂O andMeCN both buffered with 0.1% trifluoroacetic acid were used as themobile phase, HPLC conditions: UV collection 254 nm, flow rate 20mL/min, 25%→47% MeCN linear gradient over 50 minutes and then isocraticelution with 47% MeCN over 9 minutes. The HPLC fractions were combinedand lyophilized. Title compound was isolated in 35% yield (0.025 g,0.280 mmol, >95% pure) as a white fluffy solid.

Major 8a(S) Epimer:

Prep HPLC retention time: 50.40 minutes

¹H NMR (600 MHz, Methanol-d₄) δ 7.35-7.24 (m, 15H), 5.18-5.00 (m, 7H),4.32-4.14 (m, 1H), 4.23 (dd, J=9.2, 5.3 Hz, 1H), 4.12 (d, J=5.7 Hz, 1H),4.08 (dd, J=14.9, 10.1 Hz, 1H), 3.78-3.52 (m, 2H), 3.61 (d, J=10.0 Hz,1H), 3.40-3.16 (m, 2H), 3.04 (dd, J=14.7, 7.6 Hz, 1H), 2.21 (dq, J=13.4,4.6 Hz, 1H), 1.88 (ddd, J=21.9, 16.2, 8.7 Hz, 2H), 1.75-1.61 (m, 7H),1.44-1.26 (m, 2H). ¹³C NMR (151 MHz, Methanol-d₄) δ 184.18, 183.51,173.85, 169.04, 168.45, 158.71, 158.70, 138.12, 137.27, 137.24, 129.59,129.48, 129.44, 129.35, 129.32, 129.21, 129.04, 128.98, 128.83, 128.78,71.53, 71.45, 70.92, 70.85, 67.88, 67.70, 58.68, 58.03, 55.34, 53.15,52.89, 51.34, 42.94, 42.85, 36.53, 36.46, 32.00, 31.91, 29.95, 29.25,27.34, 27.31, 27.02, 26.28, 23.75, 23.72

HR-MS: (M+1)⁺=787.3675 (experimental); exact mass=787.3661 (theoretical)

IR f (cm⁻¹): 3294, 1679, 1607, 1427, 1344, 1206, 1138, 844, 802, 725,698.

[α]_(D)=+0.0150 (c=1.8, MeOH)

Minor 8a(R) Epimer:

Prep HPLC retention time: 49.25 minutes

¹H NMR (800 MHz, Methanol-d₄) δ 7.36-7.27 (m, 15H), 5.19-5.01 (m, 6H),4.67 (ddd, J=12.3, 10.1, 2.8 Hz, 1H), 4.25-4.15 (m, 2H), 4.02-3.99 (m,1H), 3.85 (ddt, J=78.5, 14.1, 7.4 Hz, 1H), 3.73 (dq, J=11.5, 3.9 Hz,1H), 3.62-3.49 (m, 2H), 3.46-3.34 (m, 2H), 3.23 (qd, J=11.9, 10.1, 7.3Hz, 1H), 2.06-2.03 (m, 1H), 1.95-1.78 (m, 3H), 1.74-1.62 (m, 6H),1.45-1.33 (m, 2H). ¹³C NMR (201 MHz. Methanol-d₄) δ 182.96, 182.21,173.97, 168.49, 158.75, 158.69, 138.14, 137.25, 129.58, 129.48, 129.44,129.33, 129.30, 129.21, 129.02, 128.99, 128.85, 128.81, 128.74, 72.70,69.91, 69.85, 67.86, 67.67, 60.47, 59.92, 55.42, 55.31, 54.63, 54.37,53.09, 43.19, 42.90, 32.45, 32.38, 31.96, 31.86, 29.97, 29.53, 27.10,27.03, 26.93, 26.32, 23.75, 23.71.

HR-MS: (M+1)⁺=787.3669 (experimental); exact mass=787.3661 (theoretical)

IR f (cm⁻¹): 3295, 1680, 1609, 434, 1345, 1206, 1140, 845, 802, 25, 698.

[α]_(D)=−0.013° (c=0.48, MeOH)

Iso-Imidazole Synthesis: One Step Rearrangement

(S)-5-(2-((3aR,8aS,Z)-5-(S)-6-(benzyloxy)-5-(((benzyloxy)carbonyl)amino)-6-oxohexyl)-2,2-dimethylhexahydro-7H-[1,3]dioxolo[4,5-c]azepin-7-ylidene)hydrazine-1-carboximidamido)-2-(((benzyloxy)carbonyl)amino)pentanoicacid (24)

Ketone 14 (0.120 g, 0.223 mmol) was dissolved in anhydrous MeOH (2.2mL), then N-amino arginine 23 (0.072 g, 0.223 mmol) was added in oneportion at room temperature. The resulting reaction mixture was stirredand monitored by LCMS. Upon complete consumption of starting material(˜2 h), the mixture was evaporated to dryness to afford an orange yellowfoam as a ˜1:1 mixture of E/Z isomers. Purification was performed usingTeledyne Isco with a reverse phase 15.5 g RediSep C18 column as thestationary phase. Water and ACN with 0.1% formic acid were used as themobile phase. Column conditions: 10% ACN for 2 column volumes (CVs),10→45% ACN over 27 CVs, 45% ACN for 5 CVs, 45→100% ACN over 2 CVs, andfinally 100% ACN over 7 CVs. Pure E and Z isomers can be isolated butisomerize upon evaporation of solvents. Thus, pure condensed product wasisolated as a monoformate salt and mixture of E/Z isomers in 69% yield(0.136 g, 0.154 mmol).

¹H NMR (400 MHz, Methanol-d₄) δ 7.38-7.26 (m, 15H), 5.17 (dd, J=19.6,12.1 Hz, 2H), 5.09 (d, J=2.0 Hz, 4H), 4.71-4.61 (m, 2H), 4.27 (dd,J=9.6, 4.9 Hz, 1H), 4.21 (dd, J=8.2, 4.7 Hz, 1H), 4.14 (d, J=13.5 Hz,1H), 3.93 (d, J=12.9 Hz, 1H), 3.78 (d, J=14.6 Hz, 1H), 3.53 (d, J=14.5Hz, 1H), 3.39-3.34 (m, 2H), 3.24-3.14 (m, 2H), 3.10 (dd, J=16.2, 4.2 Hz,1H), 2.78 (d, J=16.4 Hz, 1H), 1.99-1.86 (m, 2H), 1.81-1.70 (m, 6H),1.49-1.43 (m, 2H), 1.43 (s, 3H), 1.37 (s, 3H). ¹³C NMR (151 MHz,Methanol-d₄) δ 175.27, 173.59, 158.75, 156.11, 146.70, 138.15, 137.22,129.61, 129.49, 129.39, 129.30, 129.06, 128.77, 110.30, 74.32, 73.62,68.00, 67.72, 60.12, 58.50, 54.99, 54.75, 52.89, 42.17, 32.42, 31.89,29.94, 26.18, 25.94, 24.41, 23.81, 23.54.

HR-MS: (M+1)⁺=844.4245 (experimental); exact mass=844.4245 (theoretical)

IR f (cm⁻¹): 2362, 2336, 1674, 1456, 1343, 1201, 1138, 1065, 838, 800,22, 698, 668.

2,2,2-trifloro-1λ³-ethan-1-one,(6R,7S,Z)-4-((S)-6-(benzyloxy)-5-(((benzyloxy)carbonyl)amino)-6-oxohexyl)-2-(((S)-4-(((benzyloxy)carbonyl)amino)-4-carboxybutyl)-λ⁴-azanylidene)-6,7-dihydroxy-1,2,4,5,6,7,8,8a-octahydroimidazo[4,5-b]azepin-1-iumsalt (28)

Carbazone 25 (0.040 mg, 0.0473 mmol) was added to a dry microwave vialcharged with a stir bar and then dissolved in dry and degassed CHCl₃(0.470 mL). Then, TMSCl (0.020 mL, 0.142 mmol) was added dropwise viasyringe under argon. The vial was placed in an oil bath preheated to 95°C. The reaction was stirred at this temperature for 20 h, at which pointLCMS analysis of the resulting dark reddish-brown mixture indicatedcomplete consumption of starting material and formation of desirediso-imidazole ([M+H]⁺ 827). The reaction mixture was then treated withwater (0.010 mL) and stirred at room temperature for 1 hour. LCMSanalysis indicated full deprotection of the acetonide group ([M+H]⁺787).The reaction mixture was cooled in an ice bath and diluted with 1 Maqueous TFA (1.00 mL) and CH₂Cl₂ (0.500 mL). The resulting biphasicmixture was stirred vigorously for 5 min, then layers were allowed toseparate, the bottom organic layer drown off, and the aqueous layerre-extracted twice with CH₂Cl₂ (1.00 mL). The combined organic layerswere dried over anhydrous Na₂SO₄, filtered, and concentrated to a darkbrown residue containing a 4:1 mixture of epimers, which was separatedby preparative scale HPLC according to the procedure described above.The title compound was isolated as a TFA salt in 55% yield (0.020 g,0.0260 mmol) as a white fluffy solid.

Glucosepane Tris-Trifluoroacetate (5)

Protected glucosepane 28 (0.020 g, 0.0205 mmol) was dissolved inanhydrous MeOH (0.200 mL) and the solution was purged with nitrogen for5 min. 10% Pd/C (0.021 mg, 0.0205 mmol) was added to the vial and theresulting slurry was further purged with nitrogen (1 min) before H₂ gaswas added via a double walled balloon. Upon completion as indicated byLCMS analysis (˜6 h), the reaction mixture was purged with nitrogen (5min) and TFA (0.010 mL) was added. The crude reaction was filteredthrough a pad of celite with the aid of MeOH containing 0.1% TFA (20 mL)and the filtrate was evaporated to dryness in vacuo. The crude mixturewas purified by preparative scale HPLC.

Preparatory HPLC was performed with a SunFire Prep C18 OBD 5 □m 10×150mm reversed-phase column as the stationary phase. H₂O and MeOH bothbuffered with 0.1% trifluoroacetic acid were used as the mobile phase.HPLC conditions: UV collection 254 nm, flow rate 5 mL/min, 0% MeOHlinear gradient over 5 minutes, 0%→5% MeOH over 10 min, and 5% MeOHlinear gradient over 5 min. The run was finished with a 100% MeOH wash.The HPLC fractions were combined and lyophilized, and the title compoundwas isolated in 78% yield (0.012 g, 0.0160 mmol, >95% pure) as a whitefluffy solid.

Major Diastereomer: 8a(S)

Prep HPLC Retention Time:—13.18 min

¹H NMR (800 MHz, D₂O) δ 5.14 (dd, J=12.1, 3.2 Hz, 1H), 4.24 (s, 1H),4.09 (dd, J=15.0, 10.5 Hz, 1H), 3.90-3.83 (m, 2H), 3.80 (d, J=10.8 Hz,1H), 3.72-3.64 (m, 1H), 3.57 (dt, J=13.9, 7.1 Hz, 1H), 3.49 (td, J=6.8,2.2 Hz, minor E/Z isomer), 3.31 (t, J=6.9 Hz, 2H), 3.19-3.14 (m, 1H),2.25 (ddd, J=14.9, 7.6, 3.8 Hz, 1H), 1.99-1.81 (m, 5H), 1.79-1.62 (m,4H), 1.51-1.34 (m, 2H). ¹³C NMR (201 MHz, d₂o) δ 183.37, 182.33, 173.80,167.68, 166.83, 70.00, 69.91, 57.60, 56.89, 54.10, 52.14, 52.07, 50.28,50.13, 47.11, 42.31, 42.01, 34.79, 34.69, 30.13, 27.75, 27.66, 26.37,26.33, 25.22, 24.18, 21.77.

HR-MS: (M+1)⁺=429.2450 (experimental); exact mass=429.2456 (theoretical)

IR f (cm⁻¹): 3234, 1669, 1613, 1529, 1432, 1187, 1132, 841, 800, 723.

[α]_(D)=+0.0380 (c=1.5, MeOD)

Minor Diastereoemer: 8a(R)

Prep HPLC Retention Time: 6.21 min

¹H NMR (600 MHz, Deuterium Oxide) δ 4.84 (dd, J=12.2, 3.1 Hz, 1H), 4.75(dd, J=12.0, 3.1 Hz, 0H), 4.16 (dd, J=6.8, 2.6 Hz, 1H), 3.92 (dt,J=11.9, 3.7 Hz, 1H), 3.90-3.80 (m, 3H), 3.71 (dd, J=15.6, 6.9 Hz, 1H),3.63 (d, J=15.6 Hz, 1H), 3.52 (t, J=6.8 Hz, 0H), 3.39 (dt, J=13.8, 7.1Hz, 1H), 3.33 (t, J=6.7 Hz, 2H), 2.15 (dt, J=12.6, 3.6 Hz, 1H), 1.94(tdt, J=21.8, 11.2, 4.8 Hz, 4H), 1.85-1.62 (m, 5H), 1.52-1.34 (m, 2H).

¹³C NMR (151 MHz, D₂O) δ 182.43, 181.39, 174.46, 174.29, 167.93, 167.09,71.41, 68.97, 59.61, 58.97, 54.68, 54.49, 53.72, 52.22, 45.17, 42.54,42.23, 31.03, 30.45, 28.03, 27.97, 26.32, 25.43, 24.42, 22.84, 22.11,22.05, 22.01.

HR-MS: (M+1)⁺=429.2459 (experimental); exact mass=429.2456 (theoretical)

IR f (cm⁻¹): 2941, 1674.5, 1616, 1530, 1430, 1349, 1202, 1134, 840, 801,723.

[α]_(D)=−0.022° (c=1.23, D₂O)

Glucosepane Tris-Formate

Glucosepane tris-trifluoroacete 5 (0.010 g, 0.0139 mmol) was dissolvedin 10% aqueous formic acid (0.300 mL) and stirred at room temperaturefor 30 min. The mixture was then frozen (−78° C.) and lyophilized. Thetitle compound was isolated as a white fluffy solid in 89% yield (0.007g, 0.124 mmol).

TABLE 2 Comparison to previously published ¹H and ¹³C NMR δ(ppm) valuesfor glucosepane formate salt in D₂O⁴ δ (ppm) δ (ppm) J (Hz) Lederer OurLederer Our Lederer Our ¹H NMR Assignment Assignment ¹³C NMR AssignmentAssignment Assignment Assignment Major diastereomer: 8a(S) H_(A)-5 3.163.16 C-2 167.2 166.88 ²J_(5A, 5B) 14.9 15.0 H_(B)-5 4.09 4.09 C-3a 183.0182.34 ²J_(8A, 8B) 14.5 14.9 H-6 3.80 3.80 C-5 50.4 50.32 ²J_(1′A, 1′B)13.5 13.9 H-7 4.24 4.24 C-6 70.1 70.00 ³J_(5A, 6) 2.4 — H_(A)-8 1.881.89 C-7 69.9 69.95 ³J_(5B, 6) 10.4 10.5 H_(B)-8 2.25 2.25 C-8 34.834.79 ³J_(6, 7) 2.4 — H-8_(a) 5.14 5.14 C-8_(a) 57.6 57.60 ³J_(7, 8A) 1— H_(A)-1′ 3.59 3.57 C-1′ 52.4 52.27 ³J_(7, 8B) 4.9 3.8 H_(B)-1′ 3.653.68 C-2′ 26.4 26.46 ³J_(8A, 8a) 12.2 12.1 H₂-2′ 1.73 1.71 C-3′ 22.021.93 ³J_(8B, 8a) 2.8 3.2 H₂-3′ 1.41 1.39 C-4′ 30.3 30.49 ³J_(1′A, 2′)7.1 7.1 H₂-4′ 1.88 1.87 C-5′ 55.0 55.01 ³J_(1″, 2″) 6.8 6.9 H-5′ 3.693.70 C-6′ 175.2 175.07 H₂-1″ 3.30 3.30 C-1″ 42.2 42.11 H₂-2″ 1.73 1.70C-2″ 24.3 24.26 H₂-3″ 1.89 1.89 C-3″ 28.1 28.02 H₂-4″ 3.72 3.74 C-4″54.8 54.81 C-5″ 175.4 174.83 Minor diastereomer 8a(R) H_(A)-5 3.63 3.63C-2 167.1 167.06 ²J_(5A, 5B) 15.4 15.6 H_(B)-5 3.72 3.72 C-3a 182.0181.35 ²J_(8A, 8B) 12.6 12.6 H-6 4.16 4.16 C-5 52.2 52.20 ²J_(1′A, 1′B)14.0 13.8 H-7 3.92 3.92 C-6 69.0 68.99 ³J_(5A, 6) <1 — H_(A)-8 1.82 1.82C-7 71.4 71.42 ³J_(5B, 6) 6.6 6.8 H_(B)-8 2.15 2.15 C-8 31.0 31.00³J_(6, 7) 3.0 2.6 H-8_(a) 4.84 4.84 C-8_(a) 59.6 59.61 ³J_(7, 8A) 11.911.9 H_(A)-1′ 3.39 3.38 C-1′ 53.8 53.81 ³J_(7, 8B) 3-4 3.7 H_(B)-1′ 3.873.87 C-2′ 26.3 26.36 ³J_(8A, 8a) 11.9 12.2 H₂-2′ 1.72 1.71 C-3′ 22.022.06 ³J_(8B, 8a) 3-4 3.1 H₂-3′ 1.41 1.40 C-4′ 30.6 30.63 ³J_(1′A, 2′)7.2 7.1 H₂-4′ 1.88 1.88 C-5′ 55.2 55.23 ³J_(1″, 2″) 6.8 6.7 H-5′ 3.723.71 C-6′ 175.2 175.32 H₂-1″ 3.33 3.33 C-1″ 42.2 42.26 H₂-2″ 1.73 1.73C-2″ 24.4 24.42 H₂-3″ 1.91 1.91 C-3″ 28.2 28.16 H₂-4″ 3.76 3.76 C-4″54.9 54.96 C-5″ 175.0 174.96

Synthesis of Model Iso-Imidazoles 30:

(Z)-2-(1-benzylpiperidin-3-ylidene)-N-isobutlhydrane-1-carboxmidamide(SI-9)

SMe-semicarbazide-HI 22 (0.233 g, 1.00 mmol) was dissolved in MeOH (1mL) and iBu-NH₂ (0.100 mL, 1.00 mmol) was added drop wise via syringe.The resulting mixture was heated to 50° C. and monitored by LCMS. After1 h, LCMS indicated complete conversion to iBu-N-aminoguanidine ([M+H]⁺131.2), and to this orange solution was added commercialN-Bn-3-piperidone (0.226 g, 1.0 mmol). The resulting mixture was stirredat room temperature and monitored by LCMS. Upon full consumption ofstarting materials (˜2 h), the solvent was evaporated in vacuo to anorange residue.

Purification by column chromatography using SiO₂ as the stationary phaseand 1:1 MeCN/MeOH as the eluent yielded the title compound (SI-9) in 70%yield (0.210 g, 0.700 mmol) as orange foam.

¹H NMR (600 MHz, Methanol-d₄) δ 7.38-7.28 (m, 5H), 3.69 (s, 2H), 3.20(s, 2H), 3.10 (d, J=7.1 Hz, 2H), 2.75-2.59 (m, 2H), 2.53-2.40 (m, 2H),1.97-1.82 (m, 3H), 0.97 (d, J=6.7 Hz, 6H), ¹³C NMR (151 MHz,Methanol-d₄) δ 155.31, 155.18, 129.42, 129.04, 129.01, 128.09, 128.08,127.42, 127.27, 62.12, 62.01, 58.41, 52.40, 51.52, 51.44, 31.78, 27.95,27.93, 24.67, 24.61, 24.12, 23.04, 18.64, 18.61.

HR-MS: (M+1)⁺=302.3200 (experimental); exact mass=302.3205 (theoretical)

IR f (cm⁻¹): 3155, 2957, 1633, 1494, 1454, 1390, 1341, 1241, 1117, 747,700

4-benzyl-2-(Isobutylamino)-5,6,7,7a-tetrahydro-4H-imidazo[4,5-b]pyridin-1-iumTFA salt (30)

iBu-carbazone SI-9 (0.040 mg, 0.132 mmol) was added to a dry microwavevial charged with a stir bar and then dissolved in dry and degassedCHCl₃ (1.3 mL). Then, TMSCl (0.090 mL, 0.660 mmol) was added dropwisevia syringe under argon. The vial was capped under argon and theresulting mixture was heated in a microwave reactor to 130° C. for 15hours. The reaction mixture was then cooled to room temperature and thesolvent removed in vacuo to yield a reddish brown residue. This residuewas then taken up in CH₂Cl₂ (1 mL) and 10% aqueous TFA (1 mL). Theaqueous layer was extracted three times with CH₂Cl₂ (1 mL), and thecombined organic layers were then dried over Na₂SO₄. After filtrationand evaporation of solvent in vacuo, the resulting crude residue wassubmitted to purification.

Purification was performed on preparatory HPLC with a SunFire Prep C18OBD 10 μm 19×150 mm reversed-phase column as the stationary phase. H₂Oand MeCN both buffered with 0.1% trifluoroacetic acid were used as themobile phase. HPLC conditions: UV collection 254 nm, flow rate 20mL/min, 10%→45% MeCN linear gradient over 40 minutes. The HPLC fractionswere combined and lyophilized to give a yellow oil in 33% yield (0.017g, 0.0436 mmol)

Prep HPLC Retention Time: 24.52 min

¹H NMR (600 MHz, Methanol-d₄) δ 7.41-7.32 (m, 5H), 4.91-4.80 (m, 3H),4.76 (d, J=14.6 Hz, 1H), 4.69 (dd, J=11.6, 7.4 Hz, 1H), 4.63 (dd,J=11.6, 7.3 Hz, E/Z minor isomer), 3.52-3.38 (m, 2H), 3.37 (d. J=6.9 Hz.E/Z minor isomer), 3.10 (d, J=7.1 Hz, 2H), 2.47 (ttd, J=15.3, 7.6, 4.0Hz, 1H), 1.98-1.84 (m, 2H), 1.84-1.76 (m, 1H), 1.58-1.43 (m, 2H), 1.00(d, J=6.7 Hz, 3H), 0.97 (dd, J=6.7, 1.1 Hz, E/Z minor isomer).

¹³C NMR (151 MHz, Methanol-d₄) δ 181.51, 180.85, 170.53, 169.93, 136.36,130.01, 129.46, 129.38, 129.36, 60.37, 59.99, 54.44, 54.19, 51.51,51.15, 47.09, 46.89, 30.13, 29.48, 25.23, 25.18, 20.21, 20.18, 18.86,18.83.

¹H NMR (600 MHz, Chloroform-d) δ 10.34 (s, 1H), 9.50 (s, 1H), 7.38 (d,J=7.0 Hz, 3H), 7.30-7.27 (m, 2H), 4.79 (s, 2H), 4.42 (d, J=9.7 Hz, 1H),3.35 (t, J=6.3 Hz, 4H), 2.54 (s, 1H), 1.92 (dp, J=13.2, 6.5 Hz, 1H),1.88-1.78 (m, 2H), 1.52 (q, J=10.2 Hz, 1H), 0.96 (d, J=7.0 Hz, 6H). ¹³CNMR (151 MHz, Chloroform-d) δ 179.82, 169.53, 134.40, 129.31, 128.88,128.47, 59.07, 53.80, 50.78, 45.94, 29.05, 24.51, 20.12, 18.47.

HR-MS: (M+1)⁺=285.2065 (experimental); exact mass=285.2074 (theoretical)

IR f (cm⁻¹): 2962, 1685, 1611, 1510, 1350, 1201, 1131,801,720, 702.

(Z)-2-(1-(diethylamino)propan-2-ylidene)-N-isobutylhydrazine-1-carboximidamide(SI-10)

SMe-semicarbazide HI 22 (0.233 mg, 1.00 mmol) was dissolved in MeOH (1mL) and iBu-NH2 (0.100 mL, 1.00 mmol) was added drop wise via syringe.The resulting mixture was heated to 50° C. and monitored by LCMS. After1 h, LCMS indicated complete conversion to iBu-N-anminoguanidine([M+H]⁺131.2), and to this orange solution was added1-(diethylamino)propan-2-one (0.129 mg, 1.0 mmol). The resulting mixturewas stirred at room temperature and monitored by LCMS. Upon completionand full consumption of starting materials (˜2 h), the solvent wasevaporated in vacuo to a brown viscous oil. Purification was performedusing Teledyne Isco with a reverse phase 15.5 g RediSep C18 column asthe stationary phase. Water and ACN buffered with 0.1% FA were used asthe mobile phase. Column conditions: 0% ACN for 4 column volumes (CV)followed by 0→20% ACN over 25 CVs, 20% ACN for 5 CVs, 20→100% ACN over 2CVs, and finally 100% ACN over 5 CVs. Pure condensed product wasisolated as a monoformate salt and mixture of E/Z isomers in 58% yield(0.167 g, 0.583 mmol).

¹H NMR (600 MHz, Methanol-d₄) δ 8.44 (s, 1H), 4.06-4.00 (m, 2H), 3.26(dddd, J=17.2, 10.4, 6.9, 2.7 Hz, 4H), 3.15 (d, J=7.1 Hz, 2H), 2.05 (d,J=2.4 Hz, 3H), 1.94 (hept, J=6.7 Hz, 1H), 1.32 (qd, J=7.4, 5.9, 2.6 Hz,6H), 0.99 (d, J=6.6 Hz, 6H). ¹³C NMR (151 MHz, Methanol-d₄) δ 168.93,156.71, 50.13, 50.09, 49.77, 29.29, 20.14, 15.71, 15.66, 9.52, 9.44.

HR-MS: (M+1)⁺=242.2340 (experimental); exact mass=242.2345 (theoretical)

IR f (cm⁻¹): 2966, 1670, 1613, 1470, 1372, 1345, 1201, 1176, 1131, 830,801, 720.

5-(diethylamino)-2-(isobutylamino)-4-methyl-4H-imidazol-3-ium TFA salt(29)

iBu-carbazone SI-10 formate salt (0.038 mg, 0.132 mmol) was added to adry microwave vial charged with a stir bar and then dissolved in dry anddegassed CHCl₃ (1.3 mL). Then TMSCl (0.090 mL, 0.660 mmol) was addeddrop wise via syringe under argon. The vial was capped under argon andthe resulting mixture was heated in a microwave reactor to 130° C. for15 hours. The reaction mixture was then cooled to room temperature andsolvent removed in vacuo to yield a reddish brown residue which wassubmitted directly to purification.

Purification was performed on preparatory HPLC with a SunFire Prep C18OBD 10 μm 19×150 mm reversed-phase column as the stationary phase. H₂Oand MeCN both buffered with 0.1% trifluoroacetic acid were used as themobile phase. HPLC conditions: UV collection 254 nm, flow rate 20mL/min, 20%-60% MeCN linear gradient over 25 minutes. The HPLC fractionswere combined and lyophilized to a pale yellow residue in 12% yield(0.004 g, 0.0158 mmol).

¹H NMR (600 MHz, CDCl₃) δ 10.73 (s, 1H), 9.58 (d, J=5.7 Hz, 1H), 4.62(q, J=6.7 Hz, 1H), 3.63 (dp, J=24.9, 6.8 Hz, 2H), 3.39 (ddt, J=38.5,14.4, 7.2 Hz, 2H), 3.27 (h, J=6.7 Hz, 2H), 1.88 (hept, J=6.7 Hz, 1H),1.50 (d, J=6.7 Hz, 3H), 1.33 (t, J=7.1 Hz, 3H), 1.26 (t, J=7.1 Hz, 3H),0.93 (d, J=6.7 Hz, 6H), 13C NMR (151 MHz, CDCl₃) δ 180.56, 168.07,57.25, 50.51, 44.71, 43.71, 29.16, 20.09, 18.75, 14.10, 12.05.

HR-MS: (M+1)⁺=225.2066 (experimental); exact mass=225.2074 (theoretical)

IR f (cm⁻¹): 1682, 1602, 1210, 1137, 726.

Evidence for Conformational Exchange (E/Z Isomerization)

The interal ratios and t_(m) were used to calculate an approximate rateby the following formula:⁵

${k = {\frac{1}{t_{m}}\ln \frac{r + 1}{r - 1}}},{{{where}\mspace{14mu} r} = {( \frac{4\; X_{a}{X_{b}( {I_{aa} + I_{bb}} )}}{I_{ab} + I_{ba}} ) - ( {X_{a} - X_{b}} )^{2}}}$

X_(a)=mole fraction of conformer AX_(b)=mole fraction of conformer BI_(aa)=integral intensity for diagonal peak (conformer A)I_(bb)=integral intensity for diagonal peak (conformer B)I_(ab) and I_(ba)=integral intensity for cross-peakSample calculation with the data provided above:

I_(aa) I_(bb) I_(ab) I_(ba) X_(a) X_(b) r t_(m) (sec) k (s⁻¹) 0.16 0.330.33 0.741 0.259 1.12 1.00 2.89

FIG. 7 shows 2D NOESY with t_(m)=1000 ms.

Glucosepane pK_(a) Study

Desired pH buffers were prepared starting with commercial pH 13 buffer(containing glycine/sodium hydroxide/sodium chloride—Fluka) andtitrating freshly prepared 6 M aqueous HCl until the desired pH wasobtained.

A stock solution (1.73 μM) of glucosepane tris-TFA salt was preparedwith MilliQ water.

20 μl of stock solution were dispensed into micro-centrifuge vials andthen 60 μl of appropriate pH buffer were added. The resulting mixturewas centrifuged for 1 min and then UV spectra was acquired on 2 μlaliquots using a NanoDrop 1000 spectrophotometer (Thermo), monitoringabsorbance at 253 and 228 nm. The results are set forth in attached FIG.8.

REFERENCES—FIRST SET

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1. A method of synthesizing gluocosepane or a glucosepane derivativecomprising reacting an azepin-one compound according to the chemicalstructure (14)

where R is a C₁-C₁₂ optionally substituted hydrocarbon group or aheterocyclic group, with a semicarbazone compound according to thechemical structure:

Where X is optionally substituted S-alkyl, an optionally substitutedS-aryl, an optionally substituted S-heterocyclyl, an optionallysubstituted O-alkyl, an optionally substituted O-aryl, an optionallysubstituted O-heterocyclyl, a NR¹R² group where R¹ and R² are eachindependently H, an optionally substituted alkyl group, including analkyl group which forms an amino acid group of ornithine or lysine wherethe distal amine of the side chain of each amino acid is linked to thesemicarbazone and the amine or the amine and carboxylic groups areprotected with a protecting group, an optionally substituted aryl groupor an optionally substituted heterocyclyl, or X is an amino acid groupobtained from a D- or L-amino acid according to the chemical structure;

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline) or R³ is a side chain derived from an amino acid preferablyselected from the group consisting of alanine (methyl), arginine(propyleneguanidine), asparagine (methylenecarboxyamide), aspartic acid(ethanoic acid), cysteine (thiol, reduced or oxidized di-thiol),glutamine (ethylcarboxyamide), glutamic acid (propanoic acid), glycine(H), histidine (methyleneimidazole), isoleucine (1-methylpropane),leucine (2-methylpropane), lysine (butyleneamine), methionine(ethylmethylthioether), phenylalanine (benzyl), proline (R³ forms acyclic ring with R_(a) and the adjacent nitrogen group to form apyrrolidine group), hydroxyproline, serine (methanol), threonine(ethanol, 1-hydroxyethane), tryptophan (methyleneindole), tyrosine(methylene phenol) or valine (isopropyl) (where the R³ side chain isoptionally protected) to provide compound 24A:

where R and X are the same as above; Compound 24A is further reactedwith trimethylsilyl chloride (TMSCl) in the presence of solvent(preferably chloroform, methylene chloride) at elevated temperature(generally, above room temperature and often at the reflux temperatureof the solvent used) to provide compound 28A

or an alternative pharmaceutical salt, non-salt compound or stereoisomerthereof, Where Y′ is H and R and X are the same as above; andoptionally, deprotecting the protected compound, which can be performedin the same pot or separated prior to deprotection.
 2. The methodaccording to claim 1 wherein R is a C₁-C₁₂ optionally substitutedhydrocarbon group (preferably an optionally substituted alkyl or arylgroup) or a heterocyclic group (preferably a heteroaryl group); and X isan optionally substituted S-alkyl, O-alkyl or a NH—R¹ group where R¹ isan optionally substituted C₁-C₁₀ alkyl group, including an ornithine orlysine moiety, which contains a protecting group on the free amineand/or carboxylic acid group, wherein said reaction steps occur inmultiple steps or in a single pot where all of the steps are performedor where the compound is optionally separated prior to deprotection. 3.The method of claim 1 wherein the entire method is performed in a singlepot.
 4. The method of claim 1 wherein the method is performed in asingle pot except for the final deprotection step.
 5. The methodaccording to claim 1 wherein X is S-Me, Y′ is H and R is a C₂ or C₃alkylene group substituted with an amino group and a carboxylic acidgroup obtained from ornithine or lysine wherein said amino group and/orsaid carboxylic acid group is protected.
 6. The method according toclaim 1 wherein X is a NH—R₁ group where R₁ is a C₃ or C₄ alkylene groupsubstituted with an amino group and a carboxylic acid group obtainedfrom ornithine or lysine wherein said amino group and/or said carboxylicacid group is protected.
 7. The method according to claim 1 wherein saidcompound 28A is reacted with SiO₂ or a weak base in the presence ofaqueous solvent to convert Y′ to a OH group.
 8. The method according toclaim 7 wherein X is S-Me.
 9. The method according to claim 8 wherein Y′is OH and X is S-Me and said compound is further reacted with aprotected ornithine or lysine amino acid at the S-Me position of theimidazole to provide a compound wherein Y′ is OH and X is an ornithineor lysine moiety linked to said imidazole group by the amine in the sidechain of ornithine or lysine and the remaining amino group and/orcarboxylic acid group of said omithine or said lysine group isprotected.
 10. The method according to claim 9 wherein said OH group atY′ is reduced to a hydrogen.
 11. The method according to claim 10wherein Y is reduced a borohydride reducing agent.
 12. The methodaccording to claim 11 wherein said borohydride reducing agent isNa(OAc)₃BH).
 13. The method according to claim 9 wherein said compoundis deprotected.
 14. A method of synthesizing glucosepane or a derivativethereof comprising reacting a compound 8 according to the chemicalstructure:

with a protected lysine or ornithine derivative according to thechemical structure:BL-NH—CH₂R Where BL is a protecting group and R is an alkylene aminoacid group obtained from lysine or ornithine which is protected on thefree amine and carboxylic acid groups of R in the presence of solvent atelevated temperature in a first step, followed by exposure to atrifluoroacetic acid solution (e.g. 5%) in solvent in the presence of ahydrosilane to provide compound 10:

Exposing compound 10 to aqueous acid at elevated temperature to providea compound which spontaneously undergoes amadori rearrangement andintramolecular trapping to provide compound 13:

which is reacted with 2,2-dimethoxypropane (DMP) in the presence ofpyridinium salt (preferably pyridinium p-toluenesulfonateor PPTS) insolvent to provide compound 14,

where R is the same as above; reacting compound 14 with a semicarbazonecompound according to the chemical structure:

Where X is an optionally substituted S-alkyl (preferably, S-Me), O-alkylor a NH—R¹ group where R¹ is an optionally substituted C₁-C₁₀ alkylgroup (preferably, NHR¹ is an ornithine or lysine moiety with the distalamine of the side chain linked to the semicarbozone and the remainingamine and/or acid groups being optionally protected to form the compound24A

where R and X are the same as above; further reacting compound 24A withtrimethylsilyl chloride (TMSCl) in the presence of solvent at elevatedtemperature to provide compound 28A:

or an alternative pharmaceutical salt, non-salt compound or stereoisomerthereof, Where Y′ is H and R and X are the same as above, andoptionally, deprotecting the protected compound.
 15. The methodaccording to claim 14 wherein X in said compound 28A is S-Me, Y′ is Hand R is the same as in claim 14

Wherein said compound is reacted with SiO₂ or weak base in the presenceof aqueous solvent to provide an intermediate where Y′ is OH, and R andX are the same as above, followed by reacting the hydroxyl-containingintermediate with a protected NH-Lys (lysine) or NH-Orn (ornithine)amino acid wherein the distil amine of the side chain of the lysine orornithine is a free amine and the remaining amine or the amine andcarboxylic acid groups in the amino acid are protected to displace theS-Me group at X and substitute a protected NH-Lys or NH-Orn group,followed by reducing the hydroxyl group at Y′ to a hydrogen group whereR is the same as above and X is a protected NH-Lys or NH-Orn group. 16.(canceled)
 17. (canceled)
 18. (canceled)
 19. A method of synthesizing acompound according to the chemical structure:

or a non-salt form, alternative salt form or stereoisomer thereof WhereR is a C₂ or C₃ alkylene group substituted with an optionally protectedamine and carboxylic acid group at the distal carbon of the alkylenegroup, X is a —NHR¹ group where R¹ is a C₃ or C₄ alkylene group ofornithine or lysine containing an amine group and/or carboxylic acidgroup of said omithine or lysine being optionally protected; Comprisingreacting a compound according to the chemical structure:

Where R is a C₂ or C₃ alkyl group of omithine or lysine containing anamine group and a carboxylic group either of which or both areoptionally protected, with a semicarbazone compound according to thechemical structure:

Where X is S-methyl or —NHR¹, where R¹ is a C₃ or C₄ alkylene group ofornithine or lysine containing an amine group and/or carboxylic acidgroup of said ornithine or lysine being optionally protected to providea compound according to the chemical structure:

Where R and X are the same as above; Reacting said semicarbozonesubstituted compound above with trimethylsilyl chloride in solvent atelevated temperature to provide a compound according to the chemicalstructure

where X is S-Me or —NHR¹, and Y′ is H, or a non-salt, an alternativesalt or a stereoisomer thereof.
 20. The method according to claim 19wherein X is S-Me and said compound is reacted with SiO₂ or weak base inthe presence of aqueous solvent to convert the bridge hydrogen (Y′═H) toa hydroxyl group (Y′ is OH).
 21. The method according to claim 20wherein after said bridge hydrogen is converted to a hydroxyl group, thecompound is reacted with a protected ornithine or lysine compound andthe S-Me group is converted to a NHR¹ group, where R¹ is the same as inclaim 19, and wherein said hydroxyl group is converted to a hydrogengroup under reducing conditions using a borohydride reducing agent andsaid protecting groups are removed to provide glucosepane or aglucosepane derivative according to the chemical structure:

Where R is a C₂ or C₃ alkylene group substituted with an amine andcarboxylic acid group at the distal carbon of the alkylene group, X is a—NHR¹ group where R¹ is a C₃ or C₄ alkylene group of ornithine or lysinecontaining an amine group and a carboxylic acid group and Y′ is H, or Anon-salt form, an alternative salt form or a stereoisomer thereof. 22.(canceled)
 23. A method for synthesizing a substituted imidazole from analdehyde or ketone of the general formula 1k:

Where Z¹ is H, an optionally substituted C₁-C₁₂ hydrocarbon group, a3-20 membered heterocyclic group, a NR¹R² group, a SR¹ or OR¹ group ortogether Z¹ and Z² form an optionally substituted 5- to 7-membered ringwhich is carbocyclic or heterocyclic; Z² is H, an optionally substitutedC₁-C₁₂ hydrocarbon group, a 3-20 membered heterocyclic group or togetherZ¹ and Z² are linked to form an optionally substituted 5- to 8-memberedring which is carbocyclic or heterocyclic; R¹ and R² are eachindependently absent (with the proviso that only one of R¹ and R² may beabsent), H, an optionally substituted C₁-C₆ alkyl, alkene or alkynegroup, an optionally substituted aryl or heterocyclic group or NR¹R² isan optionally protected amino acid group where R¹ is H or a C₁-C₃ alkylgroup and R² is a group obtained from a D- or L-amino acid according tothe chemical structure:

where R³ is a side chain derived from an amino acid preferably selectedfrom the group consisting of alanine (methyl), arginine(propyleneguanidine), asparagine (methylenecarboxyamide), aspartic acid(ethanoic acid), cysteine (thiol, reduced or oxidized di-thiol),glutamine (ethylcarboxyamide), glutamic acid (propanoic acid), glycine(H), histidine (methyleneimidazole), isoleucine (1-methylpropane),leucine (2-methylpropane), lysine (butyleneamine), ornithine(propyleneamine), methionine (ethylmethylthioether), phenylalanine(benzyl), proline (R³ forms a cyclic ring with the adjacent nitrogengroup to form a pyrrolidine group), hydroxyproline, serine (methanol),threonine (ethanol, 1-hydroxyethane), tryptophan (methyleneindole),tyrosine (methylene phenol) or valine (isopropyl), where the R³ groupand/or the carboxylic acid group is optionally protected, The methodcomprising reacting a compound of formula 1i with a semicarbazonecompound of formula S1:

Where X¹ is an optionally substituted S-alkyl, an optionally substitutedS-aryl or an optionally substituted S-heterocyclyl, an optionallysubstituted O-alkyl, an optionally substituted O-aryl, an optionallysubstituted O-heterocyclyl, a NR¹R² group where R¹ and R² are eachindependently H, an optionally substituted C₁-C₁₂ alkyl group, anoptionally substituted C₁-C₆ alkyl, alkene or alkyne group, anoptionally substituted aryl or heterocyclic group, or X¹ is an aminoacid group preferably obtained from a D- or L-amino acid

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline) and R³ is a side chain derived from an amino acidpreferably selected from the group consisting of alanine (methyl),arginine (propyleneguanidine), asparagine (methylenecarboxyamide),aspartic acid (ethanoic acid), cysteine (thiol, reduced or oxidizeddi-thiol), glutamine (ethylcarboxyamide), glutamic acid (propanoicacid), glycine (H), histidine (methyleneimidazole), isoleucine(1-methylpropane), leucine (2-methylpropane), lysine (butyleneamine),methionine (ethylmethylthioether), phenylalanine (benzyl), proline (R³forms a cyclic ring with the adjacent nitrogen group to form apyrrolidine group), hydroxyproline, serine (methanol), threonine(ethanol, 1-hydroxyethane), tryptophan (methyleneindole), tyrosine(methylene phenol) or valine (isopropyl), where the R³ side chain and/orthe carboxylic acid group is optionally protected, to obtain a compoundof the formula 1i:

or a salt form (preferably a pharmaceutically acceptable salt form, orstereoisomer thereof where Z¹, Z² and X¹ are the same as above, which isthereafter reacted with trimethylsilyl chloride in solvent (e.g.chloroform) at elevated temperature to obtain the compound 1i

Where Y′ is H, and Z¹, Z² and X¹ are the same as above or ahydrochloride salt form, alternative salt form or stereoisomer thereof),Wherein said compound is optionally deprotected.
 24. (canceled) 25.(canceled)
 26. The method according to claim 23 wherein said compound 1iis reacted with SiO₂ or a weak base in aqueous solvent to convert thehydrogen at Y′ to a hydroxyl group.
 27. The method according to claim 26wherein said hydroxyl group at Y′ is converted to a hydrogen group usinga reducing agent, preferably a borohydride reducing agent. 28.(canceled)
 29. (canceled)
 30. A compound according to the chemicalstructure:

Where Z¹ is H, an optionally substituted C₁-C₁₂ hydrocarbon group, a3-20 membered heterocyclic group, a NR¹R² group, a SR¹ or OR¹ group ortogether Z¹ and Z² link to form an optionally substituted 5- to8-membered ring which ring is carbocyclic or heterocyclic, including oneor more unsaturated bonds; Z² is H, an optionally substituted C₁-C₁₂hydrocarbon group, a 3-20 membered heterocyclic group or together Z¹ andZ² are linked to form an optionally substituted 5- to 8-membered ringwhich is carbocyclic or heterocyclic; R¹ and R² are each independentlyabsent with the proviso that no more than one of R¹ and R² is absent, H,an optionally substituted C₁-C₆ alkyl, alkene or alkyne group, anoptionally substituted aryl or heterocyclic group or NR¹R² is anoptionally protected amino acid group where R¹ is H or a C₁-C₃ alkylgroup and R² is a group according to the chemical structure:

where R³ is a side chain derived from an amino acid preferably selectedfrom the group consisting of alanine (methyl), arginine(propyleneguanidine), asparagine (methylenecarboxyamide), aspartic acid(ethanoic acid), cysteine (thiol, reduced or oxidized di-thiol),glutamine (ethylcarboxyamide), glutamic acid (propanoic acid), glycine(H), histidine (methyleneimidazole), isoleucine (1-methylpropane),leucine (2-methylpropane), lysine (butyleneamine), ornithine(propyleneamine), methionine (ethylmethylthioether), phenylalanine(benzyl), proline (R³ forms a cyclic ring with the adjacent nitrogengroup to form a pyrrolidine group), hydroxyproline, serine (methanol),threonine (ethanol, 1-hydroxyethane), tryptophan (methyleneindole),tyrosine (methylene phenol) or valine (isopropyl), where said R³ groupis optionally protected; X¹ is an optionally substituted S-alkyl, anoptionally substituted S-aryl or an optionally substitutedS-heterocyclyl, an optionally substituted O-alkyl, an optionallysubstituted O-aryl, an optionally substituted O-heterocyclyl or a NR¹R²group where R¹ and R² are each independently H, an optionallysubstituted C₁-C₁₂ alkyl group, an optionally substituted aryl group oran optionally substituted heterocyclyl, or X¹ is an amino acid groupaccording to the chemical structure:

where the amine group of the amino acid is linked to the semicarbazoneand the amine group and/or the carboxylic group is optionally protectedand R_(a) is H, C₁-C₆ alkyl or alkanol or R_(a) forms a cyclic ring withR³ (proline or hydroxyproline) and R³ is a side chain derived from anamino acid preferably selected from the group consisting of alanine(methyl), arginine (propyleneguanidine), asparagine(methylenecarboxyamide), aspartic acid (ethanoic acid), cysteine (thiol,reduced or oxidized di-thiol), glutamine (ethylcarboxyamide), glutamicacid (propanoic acid), glycine (H), histidine (methyleneimidazole),isoleucine (1-methylpropane), leucine (2-methylpropane), lysine(butyleneamine), methionine (ethylmethylthioether), phenylalanine(benzyl), proline (R³ forms a cyclic ring with R_(a) and the adjacentnitrogen group to form a pyrrolidine group), hydroxyproline, serine(methanol), threonine (ethanol, 1-hydroxyethane), tryptophan(methyleneindole), tyrosine (methylene phenol) or valine (isopropyl),where the R³ side chain is optionally protected; Y′ is H or OH, or asalt form, stereoisomer, solvate or polymorph thereof.
 31. The compoundaccording to claim 30 wherein X¹ is an optionally substitutedheteroaryl, an optionally substituted S—(C₁-C₁₂) alkyl, an optionallysubstituted O—(C₁-C₁₂) alkyl or a NH—R¹ group where R¹ is an optionallysubstituted C₁-C₁₂ alkyl group.
 32. A pharmaceutical compositioncomprising an effective amount of a compound according to claim 31, incombination with a pharmaceutically acceptable carrier, additive orexcipient, optionally in further combination with an additionalbioactive agent useful in the treatment of a diabetic disorder or toinhibit or treat disorders related to the aging process
 33. (canceled)34. (canceled)
 35. (canceled)
 36. (canceled)
 37. (canceled) 38.(canceled)
 39. A compound according to a chemical structure selectedfrom the group consisting of:

where Dod is a Bis(4-methoxyphenyl)methyl group, Cbz is abenzyloxycarbonyl group, Bn is a benzyl group and TFA is atrifluoroacetate group, or a free amine, tautomer or pharmaceuticallyacceptable salt thereof.